BACTERIA  IN  RELATION  TO  PLANT  DISEASES. 


FERDINAND  COHN. 


2.  ROBERT  KOCH. 
4.  EMILE  ROUX. 


3.  LOUIS  PASTEUR. 
S.  EMILE  DUCLAUX 


IACTERIA  ix  RELATION  TO  PLANT  DISEASES 


BY 


ERWIN  F.  SMITH, 

In  charge  of  Laboratory  of  Plant  Pathology,  Office  of  Physiology  and  Pathology, 
Bureau  of  Plant  Industry,  U.  S.  Department  of  Agriculture . 


VOLUME  ONE. 


METHODS  OF  WORK  AND  GENERAL  LITERATURE  OF  BACTERIOLOGY 
EXCLUSIVE  OF  PLANT  DISEASES. 


WASHINGTON,  D.  C. : 

Published  by  the  Carnegie  Institution  of  Washington 
September,  1905. 


CARNEGIE  INSTITUTION  OF  WASHINGTON 
PUBLICATION  No.  27 


FROM    THE  PRESS   OF 

THE  HENRY   E.    WtLKENS  PRINTING  CO. 

WASHINGTON.    0.    C. 


PREFACE. 


The  subject  of  bacterial  diseases  of  plants  is  older  than  the  poured-plate  method 
of  Koch,  but  until  recently  our  knowledge  of  it  has  been  in  a  very  chaotic  state,  it 
having  been  for  the  most  part  for  twenty-five  years  a  recognized  but  uncultivated 
field.  In  recent  years,  however,  publications  on  plant  bacteria  have  multiplied, 
and  they  now  amount  to  several  hundred  titles. 

The  writer's  studies  of  the  bacteria  themselves  and  of  the  diseases  which  they 
cause,  as  distinct  from  the  literature  of  the  subject,  began  in  1893.  At  that  time 
there  was  very  little  reliable  information  on  this  subject.  The  literature  is  now 
more  extensive,  but  it  is  nowhere  gathered  together  in  one  place  and  properly  sum- 
marized. It  has  seemed,  therefore,  for  a  long  time,  that  a  work  of  the  scope  of  the 
treatise  here  presented  might  be  clarifying  and  useful  to  many  people.  There  have 
been  published,  and  are  still  appearing,  so  many  papers  on  the  subject  of  bacterial 
diseases  of  plants  by  writers  ignorant  of  bacteriological  methods  and  indifferent  to 
the  requirements  of  modern  pathological  inquiry  that  this  whole  subject  has  been 
brought  into  disrepute.  This  is  the  only  possible  explanation  of  the  fact  that  up 
to  a  very  recent  date  writers  on  pathology  and  bacteriology  have  been  telling  their 
readers  that  there  is  no  good  evidence  of  the  existence  of  any  such  diseases. 

The  following  editorial  paragraph  from  the  Botanical  Gazette,  February,  1893, 
may  be  cited  as  indicating  the  general  feeling  on  this  subject  at  that  date: 

What  is  especially  needed  at  this  stage  of  advancement  is  the  cpntinuous  and 
systematic  examination  of  the  whole  ground  by  one  or  more  well-equipped  investiga- 
tors, and  the  publication  of  a  critical  statement  of  what  may  be  safely  accepted  as 
proven.  Even  a  summarization  of  the  present  status  of  the  subject,  without  critical 
laboratory  study,  would  be  helpful,  if  well  done. 

That  this  feeling  has  become  intensified  with  the  progress  of  time  and  the 
multiplication  of  literature  is  shown  by  the  following  citation  from  the  large  Treatise 
on  Bacteriology,  by  Miquel  and  Cambier,  published  in  1902: 

The  list  of  bacteria  capable  of  attacking  the  higher  plants  increases  rapidly  from 
day  to  day  ;  but  whether  the  experiments  of  plant  pathology  offer  greater  difficulties 
than  those  of  animal  pathology,  or  whether  the  authors  who  have  undertaken  them 
have  ignored  the  multiple  resources  which  bacteriology  offers  to-day,  many  of  the 
species  described  must  be  studied  anew,  their  monography  offering  regrettable  lacunae. 
By  the  side  of  some  fruitful  and  well-conducted  labors  we  find,  unfortunately,  alto- 
gether too  many  which  must  be  done  over  entirely. 

It  was  with  the  hope  of  making  useful  discoveries  and  clearing  up  part  of 
this  contradiction  and  uncertainty  that  the  writer  began  his  study  of  this  class  of 
diseases.  His  first  effort  in  the  way  of  preparation  was  to  supplement  his  botanical 
training  with  a  knowledge  of  bacteriological  methods  which  he  obtained  from 
standard  literature  and  competent  teachers.  His  second  effort  was  to  gather 


257067 


IV  PREFACE. 

together  and  properly  digest  all  of  the  literature  relating  to  this  subject.  This 
resulted  in  the  projection  of  a  critical  review  of  the  literature,  begun  in  1896  in  the 
American  Naturalist  but  left  unfinished,  owing  to  pressure  of  research  work,  and  a 
card  catalogue  which  is  now  here  published  in  full  with  critical  remarks.  His 
third  endeavor  was  to  carefully  work  over,  in  the  laboratory,  field,  and  greenhouse, 
as  opportunity  offered,  all  of  the  so-called  bacterial  diseases  of  plants,  submitting 
each  supposed  parasite  to  all  of  the  tests  of  modern  pathology.  The  latter  has 
proved  a  far  larger  undertaking  than  was  anticipated,  the  number  of  diseases 
attributed  to  bacteria  having  increased  rapidly  in  recent  years.  It  is  expected  that 
more  than  125  diseases  will  be  treated  or  touched  upon  in  this  monograph,  many 
of  which  have  come  under  the  writer's  own  observation.  An  attempt  has  been 
made  to  cover  the  literature  of  the  whole  world  and  to  work  over  personally  every 
disease  so  far  as  material  could  be  obtained. 

The  present  volume  contains  an  "outline  of  methods  of  work,"  which  was 
written  up  in  substantially  the  same  form  four  years  ago,  in  connection  with  the 
investigations  of  the  Laboratory  of  Plant  Pathology,  Bureau  of  Plant  Industry, 
United  States  Department  of  Agriculture,  its  publication  having  been  delayed  in 
order  to  bring  the  rest  of  the  manuscript  into  suitable  shape.  The  monograph  is 
published  in  this  form  with  the  approval  of  the  Secretary  of  Agriculture. 

The  bibliography  at  the  end  of  this  volume  covers  the  general  subject  of 
bacteriology,  exclusive  of  plant  diseases,  and  is  arranged  chronologically  by  sub- 
jects. Not  every  good  paper  is  referred  to,  but  for  the  most  part  only  such  as 
have  fallen  under  the  writer's  own  observation.  It  is  believed,  however,  that  by 
consulting  these  the  student  will  soon  be  able  to  get  hold  of  the  entire  literature  of 
any  special  branch.  The  reader  who  wishes  to  keep  pace  with  the  advance  of  the 
science  should  consult  the  International  Catalogue  (R)  published  by  the  Royal 
Society  of  London. 

The  illustrations,  especially  those  dealing  with  histology,  have  been  drawn, 
with  very  few  exceptions,  under  the  direct  personal  supervision  of  the  writer,  every 
one  of  them  when  near  completion  having  been  inspected  critically  and  modified 
in  various  details  to  correspond  as  closely  as  possible  to  the  actual  object.  The 
slides  from  which  the  drawings  have  been  made  will  be  placed  on  file  in  the 
Laboratory  of  Plant  Pathology,  where  they  may  be  consulted. 

This  monograph  is  not  intended  to  take  the  place  of  ordinary  text-books  of 
bacteriology,  of  which  there  are  now  many,  but  rather  to  supplement  the  same, 
giving  information  where  they  are  silent  or  misleading.  It  is  hoped  that  it  will 
be  of  value  not  only  to  plant  pathologists,  for  whom  it  is  primarily  intended,  but 
also  to  physicians  and  animal  pathologists  for  purposes  of  comparison.  In  its 
preparation  the  writer  has  had  also  an  eye  to  the  service  of  gardeners,  fruit-growers, 
and  all  who  take  an  intelligent  interest  in  plants.  It  is  presented  with  a  keen  sense 
of  its  imperfections,  but  with  the  hope  that  it  may  at  least  serve  its  main  purpose. 
While  the  writer  has  made  every  effort  to  be  accurate  in  statement  and  just  in 
criticism,  it  is  too  much  to  hope  that  he  has  always  succeeded,  and,  therefore,  he 
desires  to  crave  pardon  in  advance  for  all  errors  of  omission  and  commission,  taking 


PREFACE.  V 

shelter  behind  Lavoisier's  well-known  and  convenient  apology:  "Man  would  never 
give  anything  to  the  public  if  he  waited  till  he  had  reached  the  goal  of  his  under- 
taking, which  is  ever  appearing  close  at  hand  and  yet  ever  slipping  farther  and 
farther  as  he  draws  nearer."  Those  who  dwell  in  the  clearer  light  of  the  next 
generation  will  build  better  than  we  have  done  and  will  scarcely  realize  how  slowly 
and  painfully  many  of  us  have  groped  about  for  what  seems  to  them  so  plain. 

In  conclusion,  I  desire  to  make  grateful  mention  of  Dr.  Theobald  Smith, 
professor  of  comparative  pathology  in  Harvard  University  Medical  School,  and  Dr. 
Veranus  A.  Moore,  professor  of  comparative  pathology  and  bacteriology  in  Cornell 
University,  each  in  turn  in  charge  of  the  animal  pathological  investigations  of  the 
Bureau  of  Animal  Industry,  United  States  Department  of  Agriculture,  at  a  time 
when  the  writer  was  beginning  his  bacteriological  studies  and  was  perplexed  in 
many  ways.  To  friendly  advice  and  helpful  suggestions  from  these  distinguished 
men  he  owes  more  than  he  can  well  repay. 

AUGUST  25,  1905. 


CONTENTS. 


OUTLINE  OF   METHODS   OF   WORK. 


Page. 

General  Remarks 3 

The  Disease 4 

Previous  Literature 6 

Geographical  Distribution 7 

Signs  of  Disease 7 

Pathological  Histology 8 

Direct-infection  Experiments 9 

The  Organism 9 

Pathogenesis 9 

Rules  of  Proof 9 

Morphology   18 

Size,  Shape,  etc 18 

Capsules  19 

Flagella  20 

Spores — Endospores,  Arthrospores 21 

Cell-unions — Zoogloeae,  Chains,  Filaments.  .  22 

Involution-forms   23 

General  Comment 23 

Physiology   25 

Motility  26 

Growth   27 

Chemotropism   27 

Reaction  to  Stains 27 

Culture-media    29 

Nutrient  Gelatin 29 

Nutrient  Agar 31 

Silicate-Jelly  36 

Solid  Vegetable  Substances 39 

Plant  Juices   (with  and  without  the  addi- 
tion of  water) , 41 

Animal  Fluids 45 

Beef-broth  45 

Milk 46 

Litmus  Milk 48 

Rice  cooked  in  Milk 48 

Loeffler's  Solidified  Blood-serum 48 

Egg-albumen    48 

Egg-yolk   49 

-Synthetic  Media  and  Other  Special  Media.  49 

Relation  to  Free  Oxygen 51 

Surface  and  Deep  Growths 51 

Fermentation-tubes    52 

Growth  in  Hydrogen,  in  Carbon  Dioxide, 

in  Vacuo,  and  in  Nitrogen 54 

Luminosity  60 


The  Organism — Continued. 
Physiology — Continued. 

Fermentation-products    60 

Alkalies  (Ammonia,  Amins,  etc.) 61 

Reducing  Powers 62 

Hydrogen  Sulphide 62 

Mercaptan  and  Other  Odors 62 

Indol,  Phenol,  Leucin,  Tyrosin,  etc 62 

Reduction  of  Nitrates,  etc 63 

Fixation  of  Free  Nitrogen,  etc 64 

Assimilation  of  Carbon  Dioxide 64 

Pigments    64 

Crystals   66 

Enzymes    66 

Sensitiveness  to  Plant  Acids 69 

Sensitiveness  to  Alkalies 69 

Effect  of  Desiccation 70 

Effect  of  Direct  Sunlight 71 

Vitality  on  Various  Media 72 

Mixed  Cultures  and  Mixed  Infections 72 

Reaction  to  Antiseptics  and  Germicides 74 

Thermal  Relations:  Maximum,  Minimum, 
and  Optimum  Temperatures  for 
Growth;  Thermal  Death-point;  Effect 

of  Freezing 75 

Other  Host-plants 87 

Pathogenic  or  Non-pathogenic  to  Animals . .  88 

Economic  Aspects. 90 

Losses  90 

Natural  Methods  of  Infection 91 

Conditions  favoring  Spread  of  the  Disease. ...  93 

Methods  of  Prevention 93 

General  Considerations 94 

Location  of  the  Laboratory 94 

Equipment  of  the  Laboratory 94 

Care  of  the  Laboratory 96 

Preparation  and  Care  of  Culture-media 97 

The  Cleaning  and  Sterilization  of  Glassware 

and  Instruments 100 

Making  and  Transference  of  Pure  Cultures. . .  103 

The  Final  Disposal  of  Infectious  Material 106 

Methods  of  Inoculation 108 

The  Keeping  of  Records 109 

The  Making  of  Collections 117 

Distilled  Water 124 


VIII 


CONTENTS. 


Page.     | 

General  Considerations — Continued. 

Microscopes  129 

Photography  and  Photomicrography 130 

Some  Milestones  in  the  Progress  of  Bacteri- 
ology    151 

Nomenclature  and  Classifications 154 

Migula's  Classification 159 

Value  of  Morphological  Characters 176 

Value  of  Cultural  Characters 178 

Undergraduate  Work 181 

A  Final  Caution 184 


Page. 

Formulae 187 

Stains: 

General  and  Miscellaneous 187 

Cleaning  Cover-glasses 189 

Flagella-staining  189 

Capsule-stains 194 

Spore-stains  194 

Non-synthetic  Culture-media 195 

Synthetic  Culture-media 197 

Miscellaneous   200 

Fixing  Fluids 202 


BIBLIOGRAPHY,  GENERAL  LITERATURE. 


I. 
II. 

III. 
IV. 


V. 

VI. 

VII. 

VIII. 

IX. 

X. 

XI. 

XII. 

XIII. 

XIV. 

XV. 


XVI. 
XVII. 
XVIII. 

XIX. 
XX. 

XXI. 

XXII. 

XXIII. 

XXIV. 

XXV. 

XXVI. 

XXVII. 

XXVIII. 


Page. 

Journals   203 

Transactions,  Beitrage,  Jahresberich- 

ten,  Festschriften,  etc 204 

Manuals  204 

Physical,  Chemical,  Zoological,  and 
Botanical  Works  of  special  use  to 

the  Plant  Pathologist 206 

Books  and   Papers  of  more  or  less 

general  interest 210 

Important    Books    and     Papers    on 

special  human  and  animal  diseases.  212 
Predisposition,   Conditions   Favoring 

Infection  or  Immunity 214 

Symbiosis,  Antagonism 214 

Carriers  of  Infection 215 

General  Morphology  of  the  Bacteria.  215 

Spores  218 

Flagella 219 

Capsules  220 

•Stains  and  Staining  Methods 221 

Morphological  and  Physiological 
Changes  due  to  Changed  Environ- 
ment    222 

Culture-media 223 

Methods  of  Work,  Apparatus,  etc.. .  226 
Special     means     of     Differentiating 

Bacteria 229 

Aerobism,  Anaerobism 230 

Fermentations,  Gas-formation,  En- 
zymes, etc 232 

Ptomaines,  Toxins,  Antitoxins,  Se- 
rums, Phagocytosis,  etc 235 

Attenuation,  Virulence 236 

Pigments,  Green  Bacteria 236 

Reduction  anil  Oxidation 239 

Nitrifying  and  Denitrifying  Organ- 
isms, Use  of  Free  Nitrogen 239 

Use  of  Free  Carbon  Dioxide 241 

Luminous  Bacteria 241 

Hydrogen  Sulphide  and  otherwise 
unclassified  By-products 242 


Page. 

XXIX.  Action  of  Light  on  Bacteria 243 

XXX.  Effect  of  Electricity 244 

XXXI.  Action     on     Bacteria    of    Roentgen 

Rays,  Radium  Rays,  etc 245 

XXXII.  Effect  of  High  Pressure  on  Bacteria. ..  245 

XXXIII.  Action  of  Heat  and  Cold  on  Bacteria.  246 

XXXIV.  Thermophilic  Bacteria 247 

XXXV.  Resistance  to  Dry  Air 248 

XXXVI.  Action  of  Acids  and  Alkalies 249 

XXXVII.  Agglutination  and  Precipitation 249 

XXXVIII.  Antiseptics  and  Germicides 250 

XXXIX.  Chemotropism,  Thermotropism,  Geo- 

tropism,  Contact-Irritation,  etc 253 

XL.  Osmotic  Pressures 254 

XLI.  Chemical  Analysis  of  Bacteria 254 

XLII.  Distribution  of  Bacteria — Geograph- 
ical and  Altitudinal 254 

XLII  I.  Soil-Organisms;  Putrefactive  Or- 
ganisms    256 

XLIV.  Vinegar-bacteria 256 

XLV.  Silage-bacteria,  Fermentation  of  To- 
bacco, of  Indigo,  Retting  of  Flax, 
of  Sisal  Hemp,  etc.,  Softening  of 

Pickles,  Sauerkraut,  etc 256 

XLVI.  Bacteria  in  Water  and  Ice;  Dung- 
bacteria  258 

XLVII.  Milk-bacteria;  Cheese-bacteria;  But- 
ter-bacteria; Meat-bacteria 259 

XLVIII.  Bacteria  in  Bread 260 

XLIX.  Iron-bacteria 261 

L.  Sulphur-bacteria 261 

LI.  Bacteria  in  Prehistoric  Times 262 

LII.  Preparation  of  Slides,  Cultures,  etc., 

for  Museums,  etc 262 

LHI.  Stock-cultures,  how  best  kept;  Vital- 
ity on  Media 263 

LIV.  Color-charts;  Nomenclature  of  Col- 
ors   263 

LV.  Photography  and  Photomicrography. .  263 

LVI.  Methods  and  Systems  of  Classification  264 

LVII.  Useful  Catalogues 265 


ILLUSTRATIONS. 


PLATES. 


Page. 

PLATE  i.  Frontispiece. 

(1)  Ferdinand  Cohn,  founder  of  mod- 

ern systematic  bacteriology.  De- 
ceased. 

(2)  Robert  Koch,  founder  of  German 

school  of  bacteriology,  director 
of  the  Institute  for  Infectious 
Diseases  at  Berlin. 

(3)  Louis  Pasteur,  founder  of  French 

school  of  bacteriology.  De- 
ceased. 

(4)  Dr.  Roux,  one  of  the  leading  spir- 

its of  the  Pasteur  Institute. 

(5)  Em.    Duclaux,    professor    in    the 

University  of  Paris  and  director 
of  the  Pasteur  Institute.  De- 
ceased. 

2.  Bacterial     Olive-knots     produced     on 

four  plants  by  needle-pricks 10 

3.  Cross-section    of    Petiole    of    Musk- 

melon,   showing  bundles   disorgan- 
ized by  Bacillus  tracheiphilus 12 

4.  Datura    metclloldes   eight   days   after 

Inoculation  with  Bacterium  solana- 
cearum  16 

5.  Zeiss    Horizontal     Photomicrographic 

Outfit 26 

6.  Arnold     Steam      Sterilizer,     Lauten- 

schlager  Dry  Oven,  Hot  Plate,  and 
Chamberland's  Autoclave 48 

7.  Hydrogen  Generator  and  Wash   Bot- 

tles in  use 56 

8.  Thermostat-room   74 

9.  Chamberland  Autoclave 84 

10.  Engine    for   furnishing   Vacuum    and 

Compressed  Air 94 


PLATE  n. 
12. 


13- 
14- 
15. 
16. 
17- 

18. 
19. 

20. 
21. 
22. 
23- 

24. 


25- 
26. 

•27. 
28. 

29. 

30. 


TEXT    FIGURES. 


Page. 

FIG.   i.  Cross-section  of  Sweet-corn  Stem  para- 
sitized by  Bacterium  Stnvarti 4 

2.  Cross-section  of  a  Raw  Carrot,  showing 

wedging  apart   of   Parenchyma   Cells 

by  Bacillus  carotovorus 5 

3.  A  Detail  from  Fig.  2 6 

4.  Turnip-root,  showing  Bacterium  camfes- 

trc  confined  to  vicinity  of  Vessels 7 

5.  Bacterium  campcstre.    A  small  portion 

of  Fig.  4  enlarged IO 


Page. 

Culture-room,  i.  e.,  place  for  making 

Cultures  of  Bacteria  in  Still  Air loj 

Movable  Hood  of  Wood  and  Glass, 
under  which  Bacteriological  Trans- 
fers may  be  made 106 

The  Reinhold-Giltay  Microtome 120 

Distilled-water  Apparatus 124 

Zeiss  Stand  lla 129 

Zeiss  Photomicrographic  Stand  Ic....   129 
Mounted  Camera  for  Enlarging,  Re- 
ducing, and  Natural-size  Work 134 

Lantern-slide  Room 144 

Black  Spot  of  the  Plum 148 

Bacterial  Disease  of  Broomcorn 150 

Bacterial  Black  Spot  of  Walnut 174 

Ditto,  Late  Stage 176 

Transmission   of   Wilt   of   Cucumber 

by  Insects 178 

Brown  Rot  of  Potato.  Natural  Infec- 
tion of  Tuber,  Artificial  Infection  of 

Stems 202 

Brown  Rot  of  Potato.     Shoots  wholly 

destroyed  by  inoculation 202 

Tomato-plant    inoculated    with   Bac.- 

terium  solanacearum 202 

Bacterial  Wilt-disease  of  Tobacco 202 

Young  Pear-shoots  blighted  by  Bacil- 
lus amylovorus 202 

Green  Pear-fruits  eight  days  after  In- 
oculation with  Bacillus  amylovorus.  202 
Quince-shoots  and  Pear-fruits  (cross- 
section)     showing     Blight    due     to 

Bacillus  amylovorus 202 

Small  Green  Apples  blighted  by  Ba- 
cillus amylovorus 202 


Page. 


FIG.  6.  Turnip-root,  showing  Bundle  occupied 
by  Bacterium  camfestre  and  the  com- 
mencement of  a  cavity;  a  later  stage 
than  Fig.  5 1 1 

7.  Cauliflower-petiole,  showing  Bundle  de- 

stroyed by  Bacterium  camfestre 12 

8.  Melon-wilt  due  to  Bacillus  tracheiphilus.     13 

9.  Cross-section  of  Bundle  of  a  Cucumber- 

stem,  showing  Bacillus  tracheiphilus 
restricted  to  the  Spiral  Vessels  and 
Three  pitted  vessels 15 


ILLUSTRATIONS. 


Page. 

FIG.  10.  Datura  metelloides  Inoculated  by  Needle- 
pricks  with  Bacterium  solanacearum. 
The  same  plant  as  in  Plate  4,  but  six 
days  later 17 

11.  (a)  Capsule   of  Organism  plated   from 

Black  Spot  of  Plum;  (b)  Viscid  Cul- 
ture-medium from  which  a  was  ob- 
tained    18 

12.  Yellow  Ooze  from  Black  Spot  of  Plum 

stained  by  ordinary  method 19 

13.  Tenuous  Threads  of  Bacillus  tracheiphi- 

lus  drawn  from  a  Muskmelon  Stem  . .     19 

14.  A  detail  from  Fig.  13,  highly  magnified.     19 

15.  Flagella  stained  from  a  pure  culture  of 

a  Bacterium  grown  in  Water  contain- 
ing a  few  drops  of  Uschinsky  Solution.  21 

16.  Beyerinck's  Drop  Bottle 21 

17.  Double  Blow  Bulb 22 

18.  Short    Form    of    Bacterium    campestre 

when  crowded 23 

19.  Long    Form    of    Bacterium    campestre 

when  grown  on  Sugar-agar 23 

20.  Hanging-drop  Culture 24 

21.  Involution-forms  of  Bacillus  tracheiphi- 

lus  24 

22.  Y-shaped  Forms  from  Root-tubercles  of 

Clover  24 

23.  Zeiss  Compensating  Ocular,  with  Screw 

or  Filar  Eye-piece  Micrometer 25 

24.  Zeiss  Upright  Photomicrographic  Cam- 

era       26 

25.  Hand-lens  for  examining  Bacterial  Cul- 

tures       27 

26.  Hand-lens  for  examining  Bacterial  Cul- 

tures, showing  another  form  of  mount.     27 

27.  Zeiss  Cover-glass  Measurer 28 

28.  Nelson's  Photographic  Gelatin 30 

29.  Agar-agar     as     received     from     Japan. 

(Slender  "Kanten") 31 

30.  Another    form    of    Agar-agar  made    in 

Japan   (Square  "Kanten") 32 

31,32.  Gelidiums  furnishing  Agar-agar...  33,34 

33.  Agar-agar  Flour  as  received  from  Euro- 

pean Manufacturers 35 

34.  Schleicher   and    Schull's    Folded    Filter 

Papers  36 

35.  Thermo-regulator  for  Blood-serum  Oven.     37 

36.  Iris-rhizome-rot    Organism    grown    on 

Sterile  Raw  Carrot 41 

37.  Tin-box  in  which  Pipettes,  Scalpels,  etc., 

may  be  sterilized 42 

38.  Fluid    Culture   showing   rise   of   Viscid 

Precipitate  when  twirled  rapidly 42 


Page. 
FIG.  39.  Platinum-iridium  Transfer  Wires 43 

40.  Simple  way  of  filtering  with  Chamber- 

land  Bougie 44 

41.  Roux    Filter    for    separating    Bacteria 

from  their  Products 45 

42.  Section  of  the  Arnold  Steam  Sterilizer, 

showing  Principle  of  Action 46 

43.  Lautenschlager  Centrifuge 47 

44.  Wire-crate  for  holding  Media  to  be  ster- 

ilized      48 

45.  Oven  for  use  in  solidifying  Blood-serum, 

etc.,  at  Temperatures  below  100°  C.  ..     49 

46.  Simple  Rack  for  holding  Fermentation 

tubes 52 

47,48,49.  Fermentation-tubes  in  actual  use. .     53 

50.  Ordinary  Kipp  Gas-generator 54 

51.  Hempel's  Burettes  for  Gas-analysis 55 

52.  Hempel's  Simple  Pipette  for  Liquid  Re- 

agents       56 

53.  Small  Novy  Jar 57 

54.  Large   Novy  Jar;   the  most  convenient 

Form  58 

55.  Simple  Device  for  growing  organisms  in 

Nitrogen  ....'. 59 

56.  Test  for  Reduction  of  Nitrates  to  Ni- 

trites       63 

57.  Crystals  formed  in  Nutrient  Agar  as  the 

Result  of  Bacterial  Growth 66 

58.  Thick-walled    Flask    for    Filtration    or 

Evaporation  in  •vacua 67 

59.  Titration-devices 68 

60.  Sodium-hydrate  Bottle 69 

61.  Effect  of   Sunlight  on    Pear-blight   Ba- 

cillus       71 

62.  Effect  of  Sunlight  on  Bean-spot  Bacte- 

rium       71 

63.  Water-bath    for    Thermal    Death-point 

Experiments 76 

64.  Roux  Metal-bar  Thermo-regulator 77 

65.  Thermometer  for  Thermal   Death-point 

Experiments    79 

66.  Leveling  Apparatus 80 

67.  Dewar     Glass     for     Experiments     with 

Liquid  Air  81 

68.  Petri-dish  Poured  Plate  inoculated  with 

a   measured    quantity   of   a    Bouillon 
Culture  of  Bacillus  tracheiphilus 82 

69.  The  same  as  Fig.  68,  but  poured  after 

Exposure  to  Liquid  Air 83 

70.  'Stomatal   Infection  by  Bacterium  pruni 

in  Green  Fruits 84 

71.  Stomatal  Infection  by  Bacterium  pruni 

in  Leaf 86 

72.  Stomatal  Infection  by  Bacterium  pruni — 

a.  Later  Stage  in  Fruit 88 


FIG.  73.  Seedling    Sweet-corn     Plant    in     Stage 

when  most  of  Infections  occur 89 

74.  Stomatal  Infection  of  Sweet-corn  Leaf 

by  Bacterium  Stcwarti 90 

75.  A  Detail  from  Fig.  74,  highly  magnified.     91 

76.  Water-pore  Infection  by  Bacterium  cam- 

pestre 92 

77.  Bacteria  from  Fig.  76,  enlarged  2,000 93 

78.  Single   Spiral  Vessel  occupied  by  Bac- 

terium campestre 93 

79.  Water-pore    Infection    in    Cabbage;    a 

later  stage  than  that  shown  in  Fig.  76.     94 

80.  Angular    Leaf-spot    of    Cotton,    Nearly 

Natural  Size 95 

81.  End    of    Vacuum-pipe    on    Laboratory- 

table  96 

82.  Portion  of  Work-table,  showing  Simple 

Apparatus  for  Distilling  Water 97 

83.  Apparatus  for  rapidly  filling  Test-tubes 

with  Measured  Portions  of  Fluid  Cul- 
ture-media      98 

84.  Can  for  holding  Culture-media 99 

85.  Wrapped  Petri  Dishes 100 

86.  Meyer's  Hypodermic  Syringe 101 

87.  Sections  through  Tooth  of  a  Cabbage- 

leaf,  showing  Entrance  of  Bacterium 
campestre 102 

88.  Green  Cucumbers  soft-rotted  by  Bacillus 

carotovorus 103 

89.  Block  for  holding  Test-tube  Cultures. ..   104 

90.  Constant   Burner,   with   Cut-off  for  re- 

ducing Size  of  Flame 105 

91.  Steel  Sewing  Needle   (Nu.  10)  set  into 

Bone-handle  and  used  for  Puncture- 
inoculations  106 

92.  Compressed-air  Tank  and  Spray-tube...   107 

93.  Atomizers  for  use  with  92 108 

94.  Hand-sprayer   for  Distribution  of   Bac- 

teria    I09 

95.  Inoculation  Cage  for  Herbaceous  Plants,  no 

96.  Labels  from  Test-tube  Cultures m 

97.  Wooden  Labels  from  Inoculated  Plants,  in 

98.  Temperature-record  Sheets 112 

99.  Nitrate-bouillon  Records 113 

100.  Sample  from  Card-catalogue,  Two-thirds 

Actual  size 114 

101.  Heading   of  Large   Sheet   for  Volumi- 

nous Abstracts 1 14 

102.  Green-cucumber   Skin,    Contents   rotted 

out  by  Bacillus  aroideae 115 

103.  Pillsbury  Slide-boxes 116 

104.  Another  Form  of  Pillsbury  Slide-box. ..   117 

105.  Small  Paraffin-oven  used  by  writer 118 


ILLUSTRATIONS. 
Page. 


...   119 

hold- 


XI 

Page. 

FIG.  106.  Infiltrated   Tissues   embedded   in    Par- 
affin in  a  Watch-glass 1 19 

107.  Infiltrated  Material  mounted  ready  to 

cut 

108.  Drawer    with    Compartments    for 

ing  embedded  material 120 

109.  Coplin's  Staining  Jar 121 

no.  Coplin's  Staining  Jar,  cross-section 121 

in.  A    Series    of    Coplin's    Staining    Jars 

Ready  for  Use 121 

112.  A  Page  from  the  Paraffin-record-book..  122 

113.  A  Mounted  Slide  of  Serial  Sections 122 

114.  A,  Rodgers  knife  for  serial  sections;  B, 

Lentz  knife  for  cutting  hard  material 
with  slant  stroke ;  C,  Torrey  knife  for 
serial  sections;  D,  Torrey  knife  for 
free-hand  sections,  a,  b,  c,  d,  end 
views  of  A,  B,  C,  D 123 

115.  Leaf -tooth  of  Cabbage  infected  by  Bac- 

terium campestre 124 

1 16, 1 17.  Details  from  Fig.  1 15 124, 125 

118.  Stomatal    Infection   of   Cotton-leaf   by 

Bacterium  mahacearum 126 

119.  The    Reinhold-Giltay    Microtome    Ar- 

ranged for  cutting  Celloidin,  etc 127 

120.  Sub-stage      Arrangement      on      Zeiss 

Stand  Ic 130 

121.  Newer  Form  of  Zeiss-Abbe  Camera 131 

122.  Zeiss  Planar  Lenses 132 

123.  Apparatus  for  Photographing  Natural 

Size 133 

124.  Swinging  Camera  for  Equal  Lighting 

of  Exposed  Object 134 

125.  Petri-dish  Poured   Plate  photographed 

by  transmitted  light 135 

126.  Green  Leaf   (Delphinium)    with   Black 

Spots;  photographed  on  a  rapid  non- 
isochromatic  plate 138 

127.  Green  Leaf   (Delphinium)   with  Black 

Spots;  photographed  on  a  slow  i so- 
chromatic  plate 139 

128.  The  Wager  Exposure-scale 141 

129.  The   Collins-Brown   Camera,   made  by 

Folmer  &  Schwing 145 

130.  Cross-level  for  use  with  Camera 146 

131.  Device  for  cutting  out  light  in  Air-shaft.  146 

132.  Side-view  of  a  Dark-room,  convenient 

for  a  few  persons 147 

133.  Top-view  of  a  Dark-room,  convenient 

for  a  few  persons 148 

134.  Side-view    of    another    Small     Dark- 

room   .• 148 

'35-  Top-view  of  a  Small  Dark-room  shown 

in  Fig.  134 149 


XII 


ILLUSTRATIONS. 


Page. 

FIG.  136.  Case    for    protecting    Squeegee-plates 

from  Dust  and  Scratches 149 

137.  Bacterium  triloculare,  Ehrenberg"s  first 

figure  166 

138.  Bacterium  triloculare,  Ehrenberg's  sec- 

ond figure 169 

139.  Bacterium  termo,  figured  by  Cohn 170 

140.  Dallinger  and  Drysdale's  conception  of 

Bacterium  termo 170 

141.  TVrmo-like      Organism      obtained      by 

throwing  Beans  into  Water 170 


Page. 

FIG.  142.  Iris-rhizome-rot;    Crowded   Agar-plate 

after  45  hours  at  25°  C 179 

143.  Iris-rhizome-rot ;  Thin  Sowing  on  Agar 

at  end  of  4  days ;  temperature  25°  C. .   180 

144.  Bacillus  aroideae  grown  on  Agar-plate 

at  37°  to  38°  C 182 

145.  Bacillus  aroideae  grown  on  Agar-plate 

at  25°  C 183 

146.  Apparatus  for  Gradual  Substitution  of 

Alcohol  for  Water  in  Tissues 184 


BACTERIA  IN  RELATION  TO  PLANT  DISEASES. 


BY  ERWIN  F.  SMITH. 


BACTERIA  IN  RELATION  TO  PLANT  DISEASES. 


BY  ERWIN  F.  SMITH. 


PART  I. — AN  OUTLINE  OF  METHODS  OF  WORK. 

GENERAL  REMARKS. 

The  following  outline  of  methods  for  the  study  of  bacterial  diseases  of  plants, 
which  are  now  in  use  in  the  Laboratory  of  Plant  Pathology,  United  States  Depart- 
ment of  Agriculture,  has  gradually  assumed  its  present  shape  as  a  result  of  the 
writer's  field,  hot-house,  and  laboratory  experiments  during  the  past  thirteen  years. 
In  nearly  the  same  shape,  so  far  as  arrangement  is  concerned,  but  in  a  less  complete 
form,  it  was  published  in  the  American  Naturalist  in  1896.* 

The  scheme  here  presented  is  entirely  practicable  and  is  believed  to  be  not  more 
extended  than  the  exigencies  of  the  case  require ;  in  the  interest  of  better  methods 
of  work  in  plant  pathology  it  is  recommended  to  all  who  contemplate  a  special 
study  of  bacterial  diseases  of  plants,  and  also  particularly  to  those  who  intend  to 
describe  and  name  species  of  bacteria,  whether  pathogenic  or  nonpatliogenic.  Those 
who  doubt  the  necessity  for  so  much  work  are  advised  to  read  procedures  recom- 
mended for  the  study  of  bacteria  by  a  committee  of  the  American  Public  Health 
Association,  and  the  earlier  paper  by  H.  Marshall  Ward  (Bibliog.,  III).f  It  would 
be  still  more  to  the  point  if  they  would  isolate  a  dozen  bacterial  organisms  from  the 
soil,  air,  or  water,  and  undertake  faithfully  to  identify  them  by  means  of  any  of  the 
older  descriptive  works,  e.  g.,  Eisenberg's  Diagnostik  or  Saccardo's  Sylloge  Fun- 
gorum,  or  even  by  such  recent  manuals  as  those  of  Sternberg,  Lehmanii  &  Neumann, 
Fliigge,  Migula,  or  Chester  (Bibliog.,  III).  Everyone  who  has  carefully  inquired 
into  the  matter  knows  that  the  brief  statement  of  the  behavior  of  an  organism  on 
nutrient  agar,  on  gelatin,  and  on  two  or  three  other  media,  with  perhaps  a  loose 
statement  of  its  color  and  size,  no  longer  constitutes  a  description  which  describes. 
Such  accounts,  of  which  there  are  a  great  many,  usually  fail  to  mention  just  those 
things  which  might  serve  to  distinguish  the  organism  from  its  fellows.  If  a  new 
species  is  not  to  be  described  so  that  it  can  be  identified  by  others,  what  then  is  the 
use  of  any  name  or  any  description  ?  The  name  will  only  serve  to  encumber  future 
synonymy  and  to  recall  the  incapacity  of  its  author. 

*The  bacterial  diseases  of  plants:     A  critical  review  of  the  present   state  of  our  knowledge, 
parts  i-vi,  Am.  Nat.,  August  and  September,  1896. 
tFor  Bibliography  see  end  of  volume. 

3 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


THE  DISEASE. 


The  line  between  disease  and  health  is  sometimes  a  very  narrow  one,  especially 
when  nothing  more  is  involved  than  some  slight  change  in  function.  The  difference, 
however,  is  very  striking  in  many  of  the  diseases  here  considered.  The  writer  has 
used  the  word  "disease"  in  the  common  acceptation  of  the  term,  meaning  thereby 


J/B 


Fig.  I.* 


any  marked  deviation  from  the  normal  functions  or  structure  of  the  plant  as  it  now 
exists,  whether  wild  or  greatly  modified  by  cultivation.  In  a  sense,  such  a  change 
as  has  taken  place  in  the  cauliflower,  the  normal  flower-shoots  of  which  have  become 

*Fic.  i. — Cross-section  of  the  upper  part  of  a  sweet-corn  stem  parasitized  by  Bacterium  Stcwarti 
(Erw.  Sm.).  The  location  of  the  bacteria  is  indicated  by  black  shading.  Most  of  .the  affected  bun- 
dles are  on  the  periphery.  The  (bacteria  'have  not  escaped  into  the  parenchyma.  Jamaica,  Long 
Island,  N.  Y.,  July  16,  1902.  The  section  was  taken  'several  feet  from  the  ground,  but  the  stem  in- 
fection undoubtedly  took  place  #hrough  one  or  more  of  the  flower  nodes.  Drawn  from  photomicro- 
graph of  a  section  stained  -with  carbol-fuchsin.  Exactly  similar  sections,  but  with  a  larger  number 
of  infected  bundles,  have  been  cut  from  stems  of  sweet --corn  plants  infected  by  the  writer  in  August, 
1902,  during  the  seedling  stage  shown  in  fig.  73. 


THE   DISEASE.  5 

compacted,  aborted,  and  enlarged  into  a  fleshy  edible  mass,  might  well  be  regarded 
as  a  diseased  condition,  but  it  is  not  so  regarded  for  the  purposes  of  this  book. 
On  the  contrary,  a  soft  rot  of  the  cauliflower  head  is  regarded  as  a  disease.  Bacterial 
diseases  of  plants  usually  involve  both  functional  and  structural  changes. 

Inasmuch  as  the  word  "  symptoms  "  has  a  subjective  as  well  as  an  objective 
connotation  in  medical  terminology,  the  writer  has  preferred  to  substitute  the  word 
"  signs  "  for  those  objective  characters  which  serve  to  distinguish  one  plant  disease 
from  another. 


Fig.  2  * 

The  student  will,  naturally,  first  turn  his  attention  to  a  careful  study  of  the 
disease.  Under  this  head  should  be  considered :  (i)  Previous  literature ;  (2) 
Geographical  distribution ;  (3)  Signs  of  the  disease  ;  (4)  Pathological  histology  ; 
(5)  Direct-infection  experiments. 


*  FIG.  2. — Cross-section  of  a  raw  carrot,  showing  wedging  apart  of  parenchyma  cells  by  Bacillus 
carotovorus  Jones;  from  paraffin-infiltrated  material.  The  carrot  was  fixed  in  strong  alcohol  72 
•hours  after  placing  on  rts  cut  surface  one  loop  of  a  fluid  culture.  The  inoculation  was  made  in  Bhe 
.middle  of  a  cross-section  of  five  whole  root,  I  cm.  thick,  placed  in  a  sterile  Petri  dish.  The  surface 
of  the  root  was  sterilized  in  mercuric  chloride  water.  This  section  was  made  several  millimeters 
l>elow  the  inoculated  surface.  A  small  portion  of  it  at  X  is  'shown  more  highly  magnified  in  fig.  3. 
This  section  was  staine<l<with  carbol-fuchsin  and  bleached  in  50  per  cent  alcohol.  Drawn  under  Zeiss 
16  nrm.  apoehromatic  objective  with  No.  4  compensating  ocular  and  the  Abbe  camera. 


6 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


In  the  present  state  of  our  knowledge  (i)  and  (2)  can  usually  be  considered 
only  after  a  very  careful  study  of  (3),  (4),  and  (5),  and  of  the  organism  itself.  They 
involve  a  knowledge  of  modern  languages,  and  a  very  considerable  familiarity  with 
scientific  literature. 

PREVIOUS  LITERATURE. 

One  of  the  first  requisites  in  a  student  is  a  knowledge  of  how  to  use  literature. 
Previous  literature  is,  however,  often  of  such  a  fragmentary  and  uncertain  sort,  as 
we  shall  see,  that  it  is  impossible  to  decide  whether  a  disease  is  actually  new  or  has 
been  written  upon  before. 


Fig.  3* 

The  literature  of  plant  diseases  will  not  be  referred  to  in  this  volume,  except 
occasionally  and  incidentally.  The  bibliography  of  this  volume  deals  only  with 
general  bacteriology — human  and  animal  diseases,  methods  of  work,  etc. 


*Pic.  3. — A  detail  from  fig.  2.  Bacillus  carotovorus  wedging  apart  cells  of  the  carrot.  Drawn 
-mostly  from  one  plane.  In  placing  the  cover-glass  a  few  of  the  bacteria  have  been  crowded  out  of 
the  intercellular  spaces  into  .parts  they  did  not  originally  occupy.  X  1,000. 


THE   DISEASE. 


GEOGRAPHICAL  DISTRIBUTION. 

Geographical  distribution  is  an  exceedingly  interesting  problem  to  many 
naturalists.  The  writer  shares  this  feeling  and  has  made  every  effort  to  determine 
it,  as  far  as  possible,  for  each  disease.  There  are,  however,  still  many  gaps  in  our 
knowledge — the  whole  subject  is  so  new,  and  information  from  all  parts  of  the 
world  is  desired.  The  inner  temperature  of  plants  conforms  nearly  or  quite  to  that 

of  the  surrounding  medium,  and 
we  might  therefore  expect,  in  some 
cases  at  least,  to  find  a  rather  more 
sharply  restricted  distribution  than 
in  diseases  of  the  warm-blooded 
animals.  From  theoretical  con- 
siderations we  should  expect  the 
distribution  of  plant  diseases  to  be 
more  like  that  of  diseases  of  fish 
and  other  cold-blooded  animals. 
Whenever  the  bacterium  is  able  to 
endure  as  wide  a  range  of  temper- 
ature as  the  host-plant,  we  should 
expect  to  find  it  as  widely  distrib- 
uted. 

SIGNS  OF  THE  DISEASE. 

Great  care  should  be  exercised 
in  the  description  of  the  physical 
signs  and  of  the  lesions  due  to  the 
parasite,  so  that  the  disease  may 
be  identified  from  these  alone,  if 
necessary.  A  great  many  cases 
should  be  examined  and  the  signs 
must  be  recorded  in  detail  and  with 
great  accuracy.  It  should  be 
remembered  that  here  is  a  frequent 
opportunity  for  error  to  creep  in, 
since  the  plant  may  be  affected  by 
two  distinct  diseases  which  have  been  confused.  Good  figures  are  always  desirable, 
but  are  not  absolutely  essential.  If  possible,  however,  photographs,  pen  or  pencil 
drawings,  and  good  water-color  sketches  should  be  secured. 

*Fic.  4. — Cross-section  of  a  turnip  root,  showing  vessels  occupied  by  Bacterium  campestre  as  the 
result  of  a  pure-culture  inoculation  by  means  of  needle-pricks  on  the  leaves.  Material  fixed  in  strong 
alcohol,  infiltrated  -with  paraffin,  cut  on  the  microtome,  stained  with  safranin-picro-nigrosin,  and 
the  differential  washing  stopped  at  just  the  right  stage.  The  bacteria  are  confined  .to  the  vessels 
and  their  immediate  vicinity.  Tihey  do  not  occur  in  the  phloem,  a  small  portion  of  which  is  shown 
at  the  top  of  the  picture.  Section  made  from  the  same  root  as  fig.  6,  but  lower,  in  the  tapering 
part.  Drawn  from  a  photomicrograph.  X  85. 


O  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

When  all  is  said,  the  signs  of  many  plant  diseases,  it  must  be  admitted,  are 
much  alike,  and  this  is  particularly  true  of  the  bacterial  soft  rots.  This  is  an  added 
reason  for  studying  them  in  each  case  as  critically  as  possible.  The  captious  reader 
might  also  remember  that  while  an  enormous  amount  of  painstaking  labor  has  been 
devoted  to  animal  pathology,  including  twenty  centuries  in  case  of  human  medicine, 
we  are  only  in  the  beginning,  so  to  speak,  of  our  knowledge  of  the  minute  pathology 
of  plant  diseases,  and  especially  of  those  due  to  bacteria. 

PATHOLOGICAL  HISTOLOGY. 

The  relation  of  the  parasite  to  the  tissues  of  the  host  should  be  studied  both  in 
fresh  material  and  in  stained  microtome  sections  made  from  material  properly  fixed 
and  infiltrated  with  paraffin.  The  organism  may  be  a  wound-parasite,  or  it  may  be 
able  to  enter  through  uninjured  parts,  i.  e.,  in  the  absence  of  visible  wounds.  Often  it 
affects  special  tissues  or  systems  of  tissues.  Sometimes  the  bacteria  are  quite  closely 
restricted  to  the  vascular  system,  forming  occlusions  (figs,  i,  4,  5,  7,  and  9).  Some- 
times they  spread  widely  in  the  intercellular  spaces  of  the  parenchyma,  forming 
numerous  cavities  (figs.  2,  3,  and  6).  Sometimes  there  are  striking  reactions  on  the 
part  of  the  host,  e.  g.,  an  enormous  multiplication  of  cells  resulting  in  cankers  or 
tumors  (plate  2).  The  habits  of  the  parasite  and  the  behavior  of  the  tissues  of  the 
host  are  best  learned  from  serial  sections.  The  student  should  not  fail  to  preserve 
(properly  labeled)  in  strong  alcohol  an  abundance  of  typical  diseased  material  for 
future  study,  exchange,  or  reference.  Stained  cover-glass  preparations  and  stained 
sections  should  also  be  mounted  in  xylol-balsam,  carefully  labeled,  and  filed  away. 
Neglect  of  these  precautions  prevents  the  experimenter  from  furnishing  the  con- 
vincing proofs  in  case  his  printed  or  oral  statements  are  called  in  question. 

As  to  the  best  methods  of  fixing  plant  material  containing  bacteria  much  remains 
to  be  learned.  The  writer  has  had  best  success  with  strong  alcohol  (90  per  cent  to 
absolute)  and  with  picric  acid  dissolved  to  saturation  in  absolute  alchohol  and  used 
boiling  hot.  In  general  the  watery  fixatives  can  not  be  used  because  they  do 
not  hold  the  bacteria  in  place ;  even  alcohol  as  strong  as  70  per  cent  allows  many 
kinds  of  bacteria  to  diffuse  out  into  the  fluid.  Boiling  absolute  alcohol  saturated 
with  mercuric  chloride  is  sometimes  useful.  The  alcohol  may  be  boiled  in  an  open 
Erlenmeyer  flask  set  on  wire  gauze  on  an  iron  tripod  over  a  small  Bunsen  flame. 
The  alcohol  is  first  brought  to  a  boil.  The  pieces  of  tissue  are  then  thrown  in  and 
allowed  to  remain  3  to  5  minutes.  It  is  better  to  divide  the  material  into  pieces 
suitable  for  embedding  before  fixing  rather  than  after.  Usually  such  a  piece  should 
not  measure  more  than  one-half  square  centimeter  or  one-half  cubic  centimeter. 
As  far  as  possible  only  fresh  material  should  be  used  for  this  purpose.  Old  material 
has  often  absorbed  air  in  quantity  sufficient  to  render  infiltration  with  paraffin  impos- 
sible or  at  least  very  difficult.  In  such  cases  infiltration  in  vacuo  will  often  render 
good  service.  The  writer  uses  a  specially  devised  air-tight  paraffin  bath  connected 
to  the  vacuum-pump.  Even  this  device  will  not  in  every  instance  insure  perfect 
infiltration. 


RULES  OF  PROOF.  9 

DIRECT-INFECTION  EXPERIMENTS. 

Direct-infection  experiments  will  frequently  separate  out  a  parasite  which  is 
overwhelmed  by  some  saprophyte  and  thus  furnish  better  material  for  plate-cultures, 
and  they  are  also  sometimes  very  useful  when  one  is  remote  from  laboratories  and 
so  situated  that  it  is  impossible  to  obtain  pure  cultures.  It  is,  however,  a  crude 
method  and  only  to  be  employed  when  more  exact  methods  can  not  be  used  or 
would  not  serve  as  well.  By  "direct  "  infection  is  meant  the  transfer  of  fluids  or 
solids  from  the  diseased  plant  directly  into  the  tissues  of  the  healthy  plant,  an  effort 
being  made  to  include  some  of  the  supposed  parasites  in  this  transfer.  It  is  a  con- 
venient expression  and  will  be  used  often  in  this  book. 


THE  ORGANISM. 

This  may  be  considered  under  three  heads — its  ability  to  produce  disease,  its 
form,  and  its  physiological  peculiarities.  Many  of  the  latter  might  equally  well  be 
denominated  cultural  characters,  and  the  pathogenic  properties  really  belong  under 
physiology,  but  are  kept  distinct  for  sake  of  convenience  and  because  they  constitute 
not  only  the  most  important  attributes  of  the  organism,  economically  speaking,  but 
also  a  distinct  and  peculiar  phase  of  the  investigation. 

PATHOGENESIS. 

What  constitutes  proof  of  the  pathogenic  nature  of  any  organism  ?  Upon  the 
ability  of  the  student  to  give  a  proper  answer  to  this  question  depends  very  largely 
his  success  or  failure  as  an  investigator.  Henle  perceived  clearly  what  was  neces- 
sary as  long  ago  as  1840,  and  Koch's  rules  are  still  fresh  in  the  minds  of  all.  There 
is  consequently  now  so  good  an  understanding  of  this  subject  among  animal  patholo- 
gists  and  professional  bacteriologists  that  if  this  book  were  designed  principally  for 
such  persons  no  comment  would  be  necessary.  A  glance,  however,  at  the  literature 
of  plant  diseases  shows  that  many  of  the  writers  on  bacterial  diseases  of  plants  have 
not  had  this  professional  training.  The  four  cardinal  requirements,  as  understood 
by  the  writer,  are  as  follows : 

RULES  OF   PROOF. 

(a)  Constant  association  of  the  organism  with  the  disease. 
(b~)  Isolation  of  the  organism  from  the  diseased  tissues  and  careful  study  of  the 
same  in  pure  cultures  on  various  media. 

(c)  Production  of  the  characteristic  signs  and  lesions  of  the  disease  by  inocu- 

lations from  pure  cultures  into  healthy  plants. 

(d)  Discovery  of  the  organism  in  the  inoculated,  diseased  plants,  re-isolation  of 

the  same,  and  growth  on  various  media  until  it  is  determined  beyond 
doubt  that  the  bacteria  in  question  are  identical  with  the  organism 
which  was  inoculated. 


10 


BACTERIA  IN  RELATION  TO  PLANT  DISEASES. 


Under  (a)  there  should  be  numerous  observations  on  many  plants,  with  very 
careful  microscopic  examination  of  stained  and  unstained  material.  The  cells  of 
many  plants  contain  granules  which  often  dance  about  so  actively  (pedesis  or 
Brownian  movement)  as  to  be  very  deceptive,  and  yet  they  are  not  bacteria.  Living 
bacteria  in  plant  tissues  can  always  be  stained  so  as  to  stand  out  distinctly  if  the 
sections  are  well  prepared  and  sufficiently  thin.  When  bacteria  occur  in  plants  as 
parasites  they  are  usually  very  abundant  in  the  vascular  system,  or  the  parenchyma, 
or  both,  and  there  is,  so  far  as  yet  known,  always  a  distinct  breaking  down  (solution) 
of  some  portion  of  the  tissues  (see  figs.  6  and  7,  and  plate  3).  If  the  pareiichymatic 
tissues  are  sound,  if  there  is  no  bacterial  ooze  on  making  sections,  if  the  vascular 


Fig.  5  * 

system  is  not  occupied,  and  if  bacteria  can  not  be  demonstrated  in  the  tissues  by 
proper  staining,  then  it  is  very  unsafe  to  infer  their  existence  from  dancing  particles, 
no  matter  how  many  may  be  visible  in  the  unstained  sections.  Moreover,  bacteria 
may  be  present  in  some  of  the  plants  and  not  in  others,  i.  e.,  not  constantly  present, 
and  so  not  the  cause  of  the  disease.  It  is  conceivable  that  they  might  also  be  present 

*Fic.  5. — Bacterium  campestre  parasitic  in  a  turnip-root  (inoculated  plant  No.  53).  This  figure 
shows  the  bacteria  crowding  out  into  the  cells  surrounding  the  reticulated  vessels.  The  Signified 
portion  of  each  vessel  is  indicated  by  fine  dots.  Material  fixed  in  strong  alcohol,  infiltrated  with 
paraffin,  cut  on  the  microtome,  stained  with  canbol-fuchsin,  and  the  excess  of  stain  removed  in 
dilute  alcohol,  section  then  dehydrated  and  mounted  in  xylol-balsam.  Drawn  from  a  photomicro- 
graph, the  contrast  'here  indicated  being  not  greater  than  that  shown  in  the  section.  X  500  circa. 


PLATE  2. 


Bacterial  olive-knots  produced  on  four  plants  by  delicate  needle-pricks. 

•ciliated  January  4,  1904.  Photographed  May  16,  1904,  nearly  natural  size.  The  organism  came  originally  from  an  olive-knot  obtained  in 
California,  where  the  disease  has  been  very  destructive  for  a  number  of  years.  A  pure  culture  obtained  from  one  of  the  California  knots  was 
inoculated  into  young  growing  olive-shoots  and  numerous  knots  resulted.  From  one  of  these,  after  about  three  months,  the  organism  was  plated 
out  and  a  subculture  from  one  of  the  colonies  was  used  to  produce  the  knots  here  shown. 


METHODS   OF    ISOLATION. 


II 


quite  constantly,  but  merely  as  followers  of  something  else.  When  possible, 
therefore,  diseased  plants  should  be  examined  for  the  suspected  pathogen,  in  large 
numbers,  in  different  years,  and  from  widely  separated  localities.  Of  course,  if  fungi 
are  also  present  they  must  likewise  be  examined  as  to  constant  occurrence  and 
pathogenic  properties. 

Under  (/;)  all  of  the  standard  nutrient  media  should  be  tried,  and  that  repeatedly, 
until  the  student  is  entirely  familiar  with  the  appearance  and  behavior  of  the 
organism.  It  is  usually  best  to  isolate  the  organism  for  experiment  from  selected 
portions  of  the  tissue  by  means  of  Esmarch  roll-cultures  or  by  the  use  of  poured 
plates  (Petri-dish  cultures),  generally  the  latter. 

Isolations  may  also  be  made  by  inserting  a  sterile  platinum  needle  or  loop  into 
the  diseased  tissue,  obtaining  therefrom  a  little  fluid,  and  drawing  this  over  the 


Fig.  6* 

surface  of  slant  agar,  gelatin,  or  potato  a  number  of  times.  This  is  an  old  method 
introduced  by  Koch  in  1881.  If  ten  or  twelve  tubes  are  used,  the  final  streaks  will 
often  consist  only  of  scattering  colonies,  from  one  or  more  of  which  the  subcultures 
may  be  made.  The  plate  method  has  the  great  advantage  of  showing  just  how 
many  kinds  of  bacteria  are  present  in  the  tissues  (provided  they  will  all  grow  in  the 
medium  used  and  under  the  conditions  of  the  experiment),  and  just  how  numerous 
they  are.  In  case  of  viscid  organisms,  or  those  forming  compact  zooglceae  in  the 

*FiG.  6. — Cross-section  of  root  of  plant  No.  53  (turnip)  parasitized  by  Bacterium  eampestre, 
showing  an  early  stage  in  the  formation  of  a  bacterial  cavity.  The  original  section  was  made  from 
material  fixed  in  alcohol,  infiltrated  with  paraffin,  stained  with  carbol-fuohsin,  and  washed  in  a  mix- 
ture of  alcohol  and  water.  Drawn  from  a  photomicrograph.  X  500. 


12 


BACTERIA   IN    RELATION    TO    PLANT    DISEASES. 


tissues,  it  is  sometimes  desirable  to  grow  them  for  a  day  in  bouillon  before  attempt- 
ing the  plate-cultures  ;  but  one  must  then  be  on  his  guard,  since  it  is  quite  possible 
by  this  method  to  start  with  enormous  numbers  of  the  right  organism  and  have  the 
bouillon  culture  filled  with  something  else  at  the  end  of  the  24  hours. 

Pure  cultures  may  also  sometimes  be  obtained  by  cutting  out  pieces  of  the 
tissue  and  throwing  them  into  tubes  of  culture  media.  This  method,  however, 
shows  little  or  nothing  as  to  the  prevalence  of  the  organism  in  the  tissues,  and  in 


Fig.  7  * 

the  hands  of  beginners  is  very  liable  to  miscarry.  If  growth  is  obtained  it  may 
indeed  have  come  from  many  organisms  of  one  sort  pervading  the  tissues  and 
causing  the  disease,  but  it  is  not  certain  that  it  did  not  result  entirely  from  one  or 

*Fic.  7. — Bundle  in  a  cauliflower-petiole  entirely  destroyed  by  Bacterium  campestre.  The  re- 
sult of  a  pure-culture  inoculation.  Plant  No.  112  inoculated  March  10,  1897,  by  needle-punctures  on 
the  blade  of  a  leaf  without  hypodermic  injection.  First  signs  of  disease  March  20.  Petiole  put  into 
alcohol  on  April  5.  Longitudinal  section.  Tissues  surrounding  .the  bundle  entirely  free  from  bac- 
teria. Section  not  made  from  the  inoculated  leaf,  but  from  the  first  leaf  that  showed  secondary 
signs.  Drawn  from  photomicrograph  of  a  paraffin  section  stained  with  carbol-fuchsin.  X  206. 


PLATE  3. 


jfB 


Cross-section  of  petiole  of  muskmelon  No.  150  attacked  by  Bacillus  tracheiphilus . 

The  bacteria  are  confined  to  the  bundles,  in  each  of  which  cavities  have  appeared.  This  section  was  taken  from  near  the  point  marked  X  on  the  inoculated 
leaf  (see  fig.  8).  The  inoculations  were  made  on  the  blade  of  the  leaf  by  means  of  delicate  needle-pricks.  The  material  was  collected  and  fixed  in 
strong  alcohol  on  the  6th  day  after  the  appearance  of  the  disease. 


METHODS   OF    ISOLATION.  13 

more  bacteria  accidentally  introduced  from  the  surface  of  the  plant,  from  one's 
clothing  or  body,  or  from  the  air ;  or  it  may  have  resulted  from  a  few  non-pathogenic 
organisms  accidentally  present  in  the  inner  tissues  of  the  plant,  particularly  in  case 
of  roots  which  have  been  dug  some  time.  It  is  therefore  much  better  for  the 
student  to  begin  with  plate  cultures.  Generally  speaking,  the  parasite  will  be  more 
easily  obtained  in  a  state  of  purity  from  plants  or  organs  of  plants  recently  attacked 
and  from  deep  tissues,  or  from  just  within  the  margin  of  advancing  diseased  areas, 
rather  than  from  near  the  surface,  or  from  parts  which  have  been  diseased  for  a 
considerable  time. 

Parts  long  affected  almost  always  contain  mixed  growths  due  to  the  multiplica- 
tion of  saprophytes  of  various  kinds.  From  such  parts  it  is  usually  much  easier  to 
obtain  the  saprophyte  than  the  parasite,  even  if  the  latter  has  not  been  entirely 
crowded  out  and  destroyed. 


Fig.  8* 

Great  care  must  be  exercised  to  avoid  introduction  of  surface  organisms  which 
might  complicate  results,  especially  if  rapid  growers.  The  easiest  and  most  satis- 
factory way,  when  the  tissues  will  admit  of  such  treatment,  is  to  sear  the  surface 
with  a  hot  knife  or  spatula  so  as  to  burn  all  surface  organisms  and  then  cut  or  dig 
through  this  sterile  surface  with  hot  or  cold  sterile  scissors,  scalpels,  forceps,  or 
needles  to  a  part  which  has  not  been  affected  by  the  heat,  from  which  some  of  the 
diseased  fluids  and  solids  may  be  removed  on  a  sterile  platinum  loop.  I  frequently 
sear  upon  sound  tissues  at  one  side  of  the  spot  from  which  I  desire  to  make  cultures 

*Fic.  8. — Muskmelon  plant  No.  150,  inoculated  with  a  pure  culture  of  Bacillus  tracheiphilus. 
The  pricked  leaf  is  on  the  left  side.  The  section  shown  in  plate  3  was  taken  from  the  point  marked 
X,  three  days  after  the  photograph  was  made  and  ten  'full  days  after  the  inoculation. 


14  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

and  then  dig  under  into  the  periphery  of  the  diseased  portion.  If  the  tissues  are 
rather  dry  the  bacteria  may  be  forced  into  the  cavity  by  careful  squeezing,  or  some 
drops  (loops)  of  sterile  water  or  beef-bouillon  may  be  introduced  into  the  cavity  and 
stirred  around  before  the  bacteria  are  removed.  If  heat  is  inadmissible,  the  speci- 
men may  be  washed  or  soaked  for  a  time  (15  seconds  to  60  minutes)  in  mercuric 
chloride  water  (1:1000)  and  the  surface  thus  freed  from  many  contaminating  organ- 
isms. Carbolic  acid  (5  per  cent  in  water)  or  lysol  (5  per  cent  in  water)  may 
also  be  used  for  sterilizing  surfaces.  Of  course  these  substances  must  be  removed 
as  far  as  possible  before  the  surface  is  broken.  This  may  be  done  to  some  extent 
by  swabbing  with  sterile  absorbent  cotton  dipped  into  sterile  water  or  by  plunging 
into  sterile  water  and  shaking.  The  disinfectants  will  be  more  certain  to  touch 
and  sterilize  every  part  of  the  surface  if  all  adhering  particles  of  air  are  driven  off 
by  first  plunging  into  alcohol  for  a  moment. 

In  case  of  bacterial  leaf-spots  the  writer  generally  obtains  satisfactory  cultures 
by  cutting  out  the  spot  and  plunging  it  for  a  few  seconds  (15  to  45)  into  1:1000 
mercuric  chloride  water,  then  rinsing  in  sterile  water  for  a  few  minutes,  crushing  and 
throwing  into  a  tube  of  bouillon  from  which  the  plates  may  be  poured  in  course  of 
an  hour,  i.  e.,  as  soon  as  the  bacteria  from  the  interior  of  the  spot  have  had  time  to 
diffuse  into  the  bouillon.  I  frequently  crush  with  a  sterile  glass  rod,  after  throwing 
the  material  into  a  tube  of  bouillon,  or  else  on  a  small  sterile  cover-glass  which  is 
then  thrown  into  the  bouillon. 

In  cases  where  heat  and  chemical  disinfectants  are  both  inadmissible  on 
account  of  danger  of  destroying  the  organisms  within  delicate  tissues,  as  in  thin 
leaves  and  other  soft  parts,  the  bacteria  or  fungus-spores  accidentally  lodged  on 
the  surface  may  be  greatly  reduced  in  number  by  gently  rubbing  all  parts  of  the 
surface  between  the  thumb  and  finger  under  distilled  water  and  then  washing  them 
in  three  or  four  successive  beakers  of  distilled  sterile  water,  the  fragments  being 
transferred  from  one  beaker  to  the  other  by  means  of  sterile  forceps.  Of  course,  the 
thumb  and  fingers  must  be  well  cleaned  in  advance  by  scrubbing  and  sometimes  by 
the  use  of  alcohol  and  corrosive  sublimate,  followed  by  sterile  distilled  water.  When 
dry,  these  washed  specimens  may  be  scraped  into,  directly  for  plate  cultures,  or  after 
the  epidermis  has  been  peeled  off  with  cold  sterile  knives  and  forceps. 

Quantitative  detenninations  may  be  made  by  grinding  up  a  given  quantity  of 
the  suspected  plant  tissue,  e.  g.,  a  cubic  centimeter  or  a  gram,  in  a  sterile  mortar 
with  clean  sterile  sand  and  10  or  20  cc.  of  beef-broth  or  sterile  water,  and  then 
making  plates  from  carefully  measured  portions  of  the  fluid,  e.g.,  from  one  2-mm. 
loop,  from  o.i  cc.,  0.5  cc.,  etc.  A  like  number  of  check  plates  made  from  equal 
portions  of  healthy  tissues  ground  under  precisely  similar  conditions  will  soon 
demonstrate  about  how  many  colonies  are  to  be  expected  per  plate  (and  what  kind) 
as  the  result  of  surface  contamination  or  air-borne  bacteria  introduced  during  the 
process  of  grinding. 

The  procedures  described  under  c  and  d  should  be  repeated  a  number  of  times 
(the  more  the  better)  and  always  with  uninoculated  plants  in  abundance  for  compari- 
son. These  control-plants  or  check-plants  must  remain  healthy.  If  they  also  become 


BEHAVIOR   OF   CHECK-PLANTS. 


diseased,  then  the  experiments  must  be  done  over  with  more  care  and  times  enough 
to  remove  all  possible  chance  of  error.  When  check-plants  become  diseased, 
especially  in  any  number,  there  is  always  room  for  grave  suspicion.  Either  the 
experimenter  has  been  grossly  careless,  assuming  that  he  used  the  right  organism  in 
his  inoculation-experiment,  or  else  he  is  working  in  a  locality  where  the  cause  of 
the  disease  is  naturally  abundant.  In  either  case,  however  well  convinced  he  him- 
self may  be,  his  readers  will  generally  have  a  lingering  suspicion  that  even  his  inocu- 
lated plants  succumbed  not  to  what  he  inserted  into  them,  but  to  some  entirely  differ- 
ent cause  naturally  present  and  overlooked  by  the  investigator.  The  remedy  for  the 

first  is  to  learn  to  use  infectious 
material  with  more  caution,  and 
for  the  second  is  to  make  the  in- 
oculation-experiments in  localities 
or  under  conditions  where  the 
plant  shall  be  less  subject  to  natu- 
ral infection. 

If  the  experiments  must  be  per- 
formed in  localities  where  the  dis- 
ease is  naturally  present,  then  a 
large  number  of  plants  must  be 
selected  for  inoculation  and  for 
control,  and  such  a  high  percent- 
age of  infections  secured  in  the 
inoculated  plants  that  the  few 
cases  occurring  naturally  in  the 
control-plants  may  be  neglected 
as  not  casting  any  doubt  on  the 
general  result.  For  example,  if, 
in  a  region  subject  to  the  given 
disease,  100  plants  were  reserved 
for  control  and  100  similar  plants 
were  inoculated,  and  out  of  this 
number  50  of  the  latter  and  40 
of  the  former  should  contract  the 


Fig.  9* 


disease,  it  is  manifest  that  no  deductions  of  any  value  could  be  made  from  the 
experiment     All   might  be  the  result  of  some  cause  totally  different  from  the 

*Fic.  9. — Gross-section  of  a  small  part  of  a  cucumber  stem,  showing  the  parasitism  of  Bacillus 
tracheiphilus  in  one  of  die  inner  bundles.  As  yet  there  is  no  bacterial  cavity,  the  bacilli  being  con- 
fined to  the  spiral  vessels  and  a  very  few  of  the  adjacent  pitted  vessels.  Material  taken  from  a  field 
near  Washington,  D.  C.,  in  1893.  Sectioned  from  paraffin.  Drawn  from  a  photomicrograph. 
X  SO.  Introduced  for  comparison  with  plate  3.  Beginning  at  the  top,  the  tissues  occur  in  the  fol- 
lowing order:  (i)  Outer  phloem,  showing  sieve  plates;  (2)  cam.bium;  (3)  immature  xylem; 
(4)  mature  xylem,  consisting  of  pitted  vessels  and  pitted  connective  tissues;  (5)  spiral  vessels  em- 
bedded in  non-lignified  living  parenchyma,  which  is  finally  destroyed  'by  the  bacteria ;  (6)  pseudo- 
cambial  layer ;  (7)  inner  phloem ;  (8)  large-celled  parenchyma  to  either  side,  separating  this  bundle 
from  its  neighbors. 


1 6  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

assumed  cause,  the  different  number  of  cases  in  the  two  groups  of  plants  being 
accidental  variations.  If,  in  such  a  locality,  only  a  very  few  plants  are  inoculated 
and  a  few  held  as  checks,  the  evidence  becomes  still  weaker  and  would  not  be  con- 
sidered entirely  conclusive  even  though  all  of  the  inoculated  plants  should  contract 
the  disease  and  all  of  the  checks  should  remain  free,  since  in  a  region  subject 
to  a  given  disease  five  or  six  healthy  plants  may  sometimes  be  found  in  prox- 
imity to  five  or  six  diseased  ones,  although  all  may  have  appeared  healthy  earlier  in 
the  season.  The  case  is  quite  different  if  out  of  100  control-plants  and  100  inocu- 
lated plants  95  per  cent  of  the  latter  and  only  2,  5,  or  10  per  cent  of  the  former 
contract  the  disease.  It  then  becomes  a  question  of  probability  which  may  be 
converted  into  reasonable  certainty  by  several  repetitions  of  the  experiment  with 
like  results.  Of  course,  the  ideal  experiment  is  one  in  which  all  the  inoculated 
plants  contract  the  disease  and  none  of  the  control-plants,  and  in  which  a  large 
number  of  plants  has  been  used  so  as  to  exclude  all  possibility  of  the  results  being 
due  to  anything  but  the  organism  used. 

Whenever  the  disease  occurs  naturally  in  the  vicinity  selected  for  the  experi- 
ments, too  much  emphasis  can  not  be  laid  on  the  necessity  of  having  numerous 
inoculated  plants  and  numerous  controls,  and  on  the  desirability  of  repetitions  of  the 
experiment  in  different  years  and  under  different  local  conditions.  It  is  important 
also  that  the  inoculated  plants  should  be  under  healthful  conditions,  /'.  e.,  under 
conditions  as  nearly  natural  as  possible.  For  example,  proper  (natural)  conditions 
would  be  much  more  nearly  attained  by  inoculating  vigorous  plants  growing  in  the 
open  air  or  in  well-kept  greenhouses  than  by  inoculating  parts  of  the  same  plants 
cut  away  from  the  stems  and  kept  under  bell-jars.  It  is  conceivable  that  inocula- 
tions which  would  succeed  very  well  under  the  conditions  last  named,  especially 
at  abnormally  high  temperatures,  might  entirely  fail  when  under  a  more  natural 
environment. 

Not  one  of  these  four  requirements  can  be  omitted  safely.  A  chain  of  evidence 
is  not  stronger  than  its  weakest  link.  Particular  stress,  therefore,  is  laid  on  being 
able  to  produce  at  will  the  characteristic  signs  and  lesions  of  the  disease  in  healthy 
plants  by  inoculation  with  pure  cultures  of  a  given  sort;  also  on  the  -re-isolation  of  the 
organism  from  the  artificially-infected  plants  after  they  have  become  diseased ;  on  the 
subsequent  proper  behavior  of  the  organism  in  nutrient  media  ;  and  on  its  ability  to 
produce  the  disease  when  again  inoculated.  This  is  the  whole  thing  in  a  nutshell. 
The  experiments  must  be  continued  until  there  is  no  doubt  whatever  as  to  the 
pathogenic  or  non-pathogenic  properties  of  the  organism.  "Almost  certainly  path- 
ogenic" always  leaves  room  for  grave  doubt  in  the  mind  of  every  thoughtful  reader. 
As  a  rule,  the  re-isolations  should  be  made  at  a  considerable  distance  from  the  point 
of  inoculation,  particularly  if  there  is  any  doubt  whatever  as  to  the  identity  of  the 
physical  signs,  since  saprophytes  have  been  known  to  live  in  plant  tissues  for  a 
considerable  number  of  weeks  near  the  place  of  inoculation,  and,  if  abundant, 
might  cause  various  disturbances  of  nutrition  without  being  the  pathogenic  organism 
sought  for.  For  example,  one  would  be  more  likely  to  obtain  the  cause  of  the 
disease  in  pure  culture  by  attempting  isolations  from  a  plant  in  the  stage  shown  in 


PLATE  4. 


Datura  metelloides  inoculated  by  needle-pricks  with  Bacterium  solanacearum  (Erw.  Sm.). 

The  items  were  pricked  at  O  and  O'  on  July  14,  1 903,  and  the  photograph  was  made  July  22.    The  first  signs  of  wilt  appeared  the  4th  day, 

About  one-third  natural  sue. 


PURE   CULTURES. 


plate  4  than  from  the  same  plant  a  week  later  (fig.  10).  One  would  also  be  more 
certain  of  pure  cultures  by  plating  from  the  interior  of  the  plant  at  A,  B,  or  C, 
rather  than  at  X  or  Y. 

The  judgment  of  experienced  bacteriologists  as  to  the  pathogenic  nature  of 


Fig.  10* 

an  organism  may  to  a  certain  extent  be  accepted  in  absence  of  full  proof,  but 
only  for  the  time  being.  Nothing  is  absolutely  certain  which  has  not  been  experi- 
mentally demonstrated. 

*Fic.  JO. — Datura  metelloides,  inoculated  by  needle-pricks  with  Bacterium  solanacearum.    The 
same  as  plate  4,  but  six  days  later,  »'.  e.,  on  July  28. 


i8 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


If  all  experimenters  in  plant  pathology,  even  in  recent  years,  had  been  careful 
to  conform  to  these  four  rules  of  practice,  the  first  three  of  which  in  essence  were 
formulated  by  Robert  Koch  as  long  ago  as  1882,  some  very  deep  chagrins  might 
have  been  avoided. 

Owing  to  insurmountable  difficulties  many  animal  pathologists,  especially  those 
who  study  human  diseases,  now  frequently  rely  on  the  first  two  rules  as  sufficient, 
but,  if  possible,  one  should  comply  also  with  c  and  d.  Plant  pathologists  are  under 
no  such  limitations,  and  should  conform  to  each  one  of  the  above-mentioned  require- 
ments, particularly  if  they  desire  their  work  to  take  high  rank  and  to  be  generally 
accepted  as  conclusive.  Material  for  plant-inoculation  experiments  is  so  cheap  and 
easily  procured  that  a  writer  who  undertakes  to  describe  a  bacterial  disease  of  plants 
has  usually  no  good  excuse  for  leaving  any  doubt  whatever  as  to  the  pathogenic 
properties  of  the  organism.  There  is  also  no  excuse  for  limiting  the  inoculations 
to  mixtures  of  bacteria  or  to  crude  material  taken  directly  from  the  diseased  plant, 
since  every  tyro  in  bacteriology  now  knows  how  to  separate  one  organism  from 
another  in  nutrient  agar  or  gelatin  by  means  of  poured  plates  or  Petri-dish  cultures. 


"';> 


Fig.  11* 

MORPHOLOGY. 
SIZE,  SHAPE,  ETC. 

The  smallest  observed  bacteria  are  only  a  small  fraction  of  a  micron  in  diameter. 
Migula  states  that  the  stained  rods  of  Ps.  indigofera  (Voges)  Mig.  from  colonies  36 
hours  old  measured  only  0.18  by  0.06  micron.  Bacillus  denitrificans  (Amp.  &  Gar.) 
Mig.  is  also  a  very  small  rod — i.o  to  1.5  by  o.i  to  0.3  micron,  according  to  Migula. 
Micrococcus  progrediens  Schroter  is  said  to  be  only  0.15  micron  in  diameter.  The 
organism  of  peri-pneumonia  isolated  by  Nocard  &  Roux  is  more  minute.  It  is 
probable  also  that  still  smaller  organisms  occur,  i.  <?.,  so  small  as  to  be  invisible  under 
the  highest  magnifications.  In  this  way  are  interpreted  the  results  obtained  by 
animal  pathologists  in  the  foot-and-mouth  disease  and  in  some  other  diseases. 
Photographs  with  ultra  violet  light  may  in  the  end  render  some  service  here.  The 

*Fic.  ii. — a,  Capsule  of  organism  obtained  from  black  spot  of  the  plum.  Bacteria  grown  in 
Uschinsky's  solution  and  stained  by  Ribbert's  method ;  b,  ropy  Uschinsky  solution  from  which 
a  was  made. 


MORPHOLOGY. 


largest  bacteria  are  several  thousand  times  as  bulky  as  the  smallest.  Errera  has 
described  a  spirillum  the  largest  specimens  of  which  measured  23  to  28  by  3  to  3.4 
micra  ('02,  Errera,  Bibliog.,  X),  and  Schaudinn  has  described  a  bacillus  the  largest 
forms  of  which  are  24  to  80  by  3  to  6  micra  ('02,  Schaudinn,  Bibliog.,  XI). 

In  shape  the  bacteria  vary  according  to  genera  and  species  and  sometimes 
within  the  limits  of  the  species,  from  globose  cells  or  very  short  straight  rods,  through 
curved  forms  or  spirals,  to  filaments  which  are  many 
times  the  diameter  of  the  organism.  To  what  ex- 
tent does  form  vary  under  changed  conditions  ?  With 
the  eye-piece  micrometer  make  careful  measure- 
ments of  unstained  organisms  taken  from  the  host- 
plant  and  from  cultures  of  various  ages  and  kinds. 
There  is  frequently  considerable  variability  in  the  size 
of  individuals  of  the  same  species.  Is  the  breadth 
more  constant  than  the  length?  Does  the  size  or  Fig.  12.* 

shape  as  observed  in  the  plant  differ  from  that  observed  on  culture  media?  How  does 
the  living  organism  differ  in  size  and  general  appearance  from  the  dead,  stained  one? 

CAPSULES. 

The  presence  of  capsules  may  be  suspected  whenever  a  bacterial 
growth  becomes  viscid.     They  are  often  difficult    to  see  because  their 
,t  index  of  refraction  is  so  nearlv 

^  * 

v     '      ,\  .  -°» mm  .  that  of  the  fluid  in  which  they 

'         *"^   <i          ^  are  usually  examined.     In  ex- 

amining unstained  material  the 
field  should  be  illuminated  with 
*j      a  narrow  pencil  of  rays,  and  the 
j'      effect  of  illumination  with  ob- 
lique   light    should    be    tried. 
Several    methods    of    contrast 
staining  are  in  use.      By  one 
method  the  capsule  remains  un- 
m      stained  or  nearly  so,  while  the 
central  portion  of  the  bacterium 
and  the  slime  lying  on  the  cover 
between  the  bacteria  stain  more 
or    less    deeply.      By    another 
method  which    has   been   spe- 


'•• 


r 


•>-*•  i? 


V'-'-'-  »"- 

*  }          ^L 


Fig.  I3.t 


Fig.  144 


*Fic.  12. — A  portion  of  the  yellow  ooze  from  the  black  spot  of  the  plum,  stained  by  ordinary 
methods.  X  2,000. 

tFic.  13. — Cobwebby,  sticky  threads  of  Bacillus  tracheiphilus  drawn  from  the  cut  end  of  a 
muskmelon  stem,  arranged  on  a  slide  and  stained  with  carbol-fuchsin.  About  three  times  natural 
size.  Buzzards  Bay,  Mass.,  Oct.  8,  1903.  Fig.  14  was  drawn  from  the  left-hand  thread  at  the 
point  marked  X. 

JFic.  14. — Bacillus  trachciphilus  Erw.  Sm.  A  portion  of  one  of  *he  threads  shown  in  fig.  13. 
T*he  arrow  indicates  the  direction  of  the  thread,  which  was  extremely  tenacious.  The  distance  be- 
tween the  bacterial  rods  indicates  very  clearly  the  extreme  viscosity  of  iBhe  unstained  substance 
lying  between  them  and  holding  them  together.  X  1,000. 


2O  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

cially  commended  by  Dr.  Welch  ('92,  Bibliog.,  XIII),  the  capsule  is  also  stained,  but 
remains  distinctly  paler  than  the  body  of  the  bacterium.  They  may  also  be  counter- 
stained,  as  in  Muir's  method  or  Moore's  method.  Well-defined  capsules  are  shown 
in  fig.  1 1 a.  This  may  be  compared  with  fig.  12,  in  which  the  same  organism  is 
shown  without  capsules.  Fig.  nt>  shows  the  extreme  viscidity  of  a  culture  due  to 
the  formation  and  deliquescence  of  capsules.  Fig.  13  shows  the  tenuous  threads 
into  which  Bacillus  tracheiphilus  may  be  drawn  as  it  oozes  from  the  cut  stems  of 
cucurbits.  Fig.  14  is  a  detail  from  the  same  more  highly  magnified,  the  viscid  con- 
necting substance  being  unstained. 

FLAGELLA. 

Ehrenberg  was  the  first  to  describe  flagella  on  bacteria  (Bacterium  triloculare, 
1838).  Nothing  more  was  done  until  1872,  when  Colin  discovered  them  on  Spi- 
rillum vohitans.  In  1875  Dallinger  &  Drysdale  saw  and  figured  them  on  Bacterium 
tcrmo.  In  1875  Warming  determined  their  existence  on  Vibrio  rugula  and  Spi- 
rillum tmdula.  In  1877  Koch  demonstrated  their  existence  on  a  number  of  species 
by  the  use  of  stains.  In  1878  Dallinger,  using  unstained  material,  saw  them  many 
times  on  Bacterium  termo  and  also  on  Spirillum  volutans.  After  1879  110  one 
appears  to  have  disputed  their  existence.  In  1890  Messea  proposed  to  divide  the 
flagellate  bacteria  into  four  large  groups,  monotrichiate,  lophotrichiate,  amphitri- 
chiate,  and  peritrichiate.  In  1895  Fischer  used  the  flagella  as  marks  to  distinguish 
subfamilies.  In  the  previous  year  Migula  used  their  number  and  mode  of  attachment 
as  a  means  of  distinguishing  genera. 

The  staining  of  flagella  has  now  become  a  regular  part  of  laboratory  work. 
Their  number  and  position  on  the  body  wall  should  be  determined,  if  possible,  in 
case  of  each  species  studied.  This  is  sometimes  quite  easy  and  at  other  times  very 
difficult.  It  should  also  be  determined  whether  the  flagella  are  fugitive  or  persistent. 

Flagella  may  be  stained  from  young  agar  cultures.  Bouillon  cultures  are  to  be 
avoided  because  of  the  intense  ground  stain.  Some  kinds  may  be  stained  readily 
from  cultures  grown  for  some  days  in  a  very  dilute  Uschinsky's  solution — i  to  3 
drops  in  10  cc.  of  distilled  water  (fig.  15).  The  flagella  of  some  bacteria  are  stained 
readily,  those  of  others  only  with  great  difficulty.  Many  sorts  seem  inclined  to 
throw  off  their  flagella  when  transferred  from  agar  to  water.  The  cover-glasses 
must  be  clean.  When  cleaned  ready  for  use  seize  with  the  forceps  and  pass  them 
three  times  through  the  upper  part  of  the  Bunsen  flame,  with  a  considerable  interval 
between  each  flaming,  to  avoid  cracking.  Use  a  minim  quantity  of  the  culture 
stirred  in  a  big  drop  of  water,  or  even  in  2  to  10  cc.  of  water  in  a  watch  glass  or 
test  tube.  Give  the  bacteria  time  to  diffuse  by  waiting  half  an  hour  or  more.  Take 
the  cover  between  the  thumb  and  finger  of  the  left  hand,  touch  the  end  centimeter 
of  a  platinum  needle  to  the  water  containing  the  bacteria,  and  sweep  it  deftly  across 
the  cover  glass.  In  this  way  the  fluid  is  spread  in  a  very  thin  sheet  over  nearly  the 
whole  surface  of  the  cover  and  is  dry  almost  at  once,  with  the  bacteria  well  separated. 
If  the  fluid  will  not  spread,  then  the  cover  is  not  clean  and  should  be  discarded. 
The  bacterial  sheet  may  be  mordanted  and  stained  at  once,  or  first  fixed  by  gentle  heat. 
To  avoid  scorching,  the  cover  should  be  held  between  thumb  and  finger  when  it  is 
passed  rapidly  through  the  flame.  Beginners  usually  burn  the  bacterial  layer. 


STAINING   OF   FLAGELLA. 


21 


Smeary  dark  lines  aud  other  deceptive  artefacts  must  not  be  mistaken  for  the 
flagella.  The  following  methods  have  been  tried  by  the  writer  and  have  given  good 
results,  but  none  can  be  depended  upon  always,  and  much  time  and  patience  are 
sometimes  required  to  get  good  preparations  of  a  refractory  organism  :  Fischer's 

modification  of  Loeffler's 
stain ;  Moore's  modifica- 
tion of  Loeffler's  stain  ; 
Van  Ermengem's  nitrate 
?  \J  /  \ \.  \  of  silver  method ;  L6  wit's 

S  \   copper-sulphate  fuchsin 

Fi«-  15-*  mordant,  followed  by  Ehr- 

lich's  anilin-water  gentian  violet.      (For  other  methods  consult   "  Formulae  "  and 
"  Bibliography  of  General  Literature,"  XII.) 

In  connection  with  flagella-staining  a  white  porcelain  tray,  such  as  photogra- 
phers use,  will  be  found  very  convenient  for  washing,  and  also  the  double  blow-bulb 
shown  in  fig.  1 7.  This  should  be  attached  to  a  wash-bottle,  such  as  that  shown  in 
fig.  16.  This  will  deliver  a  small  stream,  veiy  good  for  washing  excess  of  mordant 
and  stain  from  the  covers.  To  furnish  a  steady  stream  the  bulb  has  to  be  compressed 
only  about  once  a  minute.  The  flask  used  for  this  purpose  should  hold  a  liter. 

SPORES — ENDOSPORES,  ARTHROSPORES. 

Do  arthrospores  really  occur?  If  so,  in  what  respect  do  they  differ  from  the 
ordinary  vegetative  rods?  Test  spores  for  resistance  to  high  temperatures  in  the 

water  bath  and  to  steam  heat;  study  germination  in 
hanging  drops.  Do  the  spores  require  a  period  of  rest 
or  refuse  to  germinate  except  in  special  media?  The 
suspected  existence  of  spores  may  be  definitely  settled 
by  seeing  the  problematic  bodies  germinate.  In  the 
absence  of  such  proof,  considerable  certainty  may  be 
reached  by  a  combination  of  two  methods:  (i)  the  use 
of  watery  basic  anilin  stains,  and  (2)  the  use  of  moist 
heat.  If  at  room  temperatures  the  glistening  bodies 
refuse  to  take  the  simple  stains  even  on  long  exposure 
and  at  the  same  time  are  very  resistant  to  steam  heat 
or  to  hot  water,  i.  e.,  much  more  so  than  the  ordi- 
nary vegetative  rods,  it  may  be  assumed  that  they  are 
spores.  If,  on  the  contrary,  they  are  destroyed  by  tem- 
peratures only  slightly  above  the  recorded  thermal  death- 
point  of  the  vegetative  rods,  it  must  not  be  assumed 
that  they  are  spores,  no  matter  how  they  behave  toward 


Fig.  16.t 


*Fic.  15. — Flagella  of  yellow  organism  plated  '£rom  black  spot  of  plum.  Stained  from  culture 
grown  in  10  cc.  distilled  water  containing  a  few  drops  of  Usohinsky's  solution.  X  1,000. 

tFic.  16. — Beyerinck's  drop-bottle.  The  size  and  number  of  drops  in  a  given  time  are  regulated 
by  sliding  the  bent  tube  through  the  cork.  It  is  very  convenient  to  have  tiiis  flask  on  die  microscope 
table.  By  a  minim  infection  of  the  fluid  it  may  also  be  arranged  so  that  each  drop  shall  deliver  a 
single  spore  or  bacterium  for  ihanging-drop  studies.  About  two-fifths  natural  size. 


22  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

stains,  unless  they  can  be  made  to  germinate.  Many  of  the  older  identifications  of 
spores  are  untrustworthy.  Alfred  Fischer  has  shown  that  many  of  these  determina- 
tions rested  on  plasmolysis  of  the  rods,  i.  e.,  on  misinterpretations.  Omelianski 
reports  finding  an  oval  spore  which  stains  readily  with  ordinary  anilin  stains.  This 
occurs  in  a  rather  large  bacillus  accompanying  his  hydrogen  cellulose  ferment.  Dan- 
napple  reports  finding  spores  which  are  very  sensitive  to  heat  ('99,  Bibliog.,  XXXIII). 
Usually  only  one  endospore  occurs  in  each  cell,  but  Kern  ('81,  Bibliog.,  VIII),  and 
Schaudinn  ('02,  Bibliog.,  XI)  have  found  bacteria  with  two  in  each  cell.  Excellent 
directions  for  the  study  of  spores  are  given  in  Part  I  of  Migula's  System  der  Bakterien 
(see  especially  the  second  paragraph  on  p.  209). 

CBLL-UNIONS— ZOOGI.CE^,  CHAINS,  FILAMENTS. 

In  some  media  bacteria  are  much  inclined  to  separate  after  division ;  in  others 
they  remain  attached  in  various  ways.  The  most  common  method  of  union  is  an 
irregular  clumping,  which  in  fluids  gives  rise  to  a  fine  or  coarse  flocculence.  Such 
unions  also  occur  on  solid  media  and  may  be  designated  zoogloese,  or  pseudo- 
zoogl&a,  if  one  prefers  to  retain  zooglosae  for  the  more  intimately  fused  and  com- 
pacter  gelatinous  unions.  Sometimes  the  organisms  remain  attached  end  to  end. 
Where  the  segmentation  is  distinct,  such  unions  are  designated  chains.  When 
very  long  and  with  obscure  segmentation,  they  may  be  called  filaments.  Is 
there  any  true  branching?  What  especial  conditions  of  the  culture  medium  favor 
the  formation  of  zoogloese,  of  chains,  and  of  filaments?  Many  bacteria  form 
zooglcese,  chains,  or  very  long  filaments  under  certain  conditions,  while  under  other 


Fig.  17* 

conditions  they  remain  as  very  short,  straight  rods.  (Compare  figs.  18  and  19.) 
As  in  case  of  involution  forms  unfavorable  cultural  conditions  (thermal,  nutrient, 
etc.)  appear  to  have  much  to  do  with  their  appearance. 

The  growth  of  bacteria  may  be  studied  in  hanging  drops  of  bouillon,  etc.  Hol- 
low-ground slides  (fig.  20)  should  be  used  for  this  purpose,  rather  than  ring-cells, 
especially  with  high  powers.  Hill's  hanging-block  method  is  also  serviceable 
('02,  Bibliog.,  XVII). 

*Pic.  17. — Double  blow-bulb  for  attachment  to  drop-bottle  shown  in  fig.  16.  By  use  of  this  de- 
vice one  obtains  with  a  minimum  of  pumping  a  constant  small  stream  of  water  very  suitable  for 
washing  stained  covers,  etc.  Made  by  Emil  Greiner.  It  is  best  used  with  a  larger  flask  than  that 
shown  in  fig.  16.  Bulbs  which  have  been  long  in  stock  should  not  be  purchased,  as  the  rubber  de- 
teriorates rapidly. 


MORPHOLOGY. 


INVOLUTION   FORMS. 

Under  this  name  we  designate  swollen  and  distorted  forms  common  in  old 

cultures  (fig.  2 1).  Under  what  conditions  do  they 
occur  ?  Are  they  living  or  dead  ?  Isolate  in 
hanging  drops  of  bouillon  and  determine  whether 
they  are  stages  in  development  or  only  degenera- 
tion forms.  Are  Y-shaped  or  branched  forms 
such  as  occur  in  old  cultures  of  B.  tuberculosis 
Koch,  and  in  the  root-tubercles  of  clover  (fig.  22) 
to  be  considered  as  involution  forms  ?  Are  such 
organisms  fungi  or  bacteria  ?  Branching  forms 
have  been  detected  by  many  observers.  (Consult 
numerous  citations  in  the  Bibliography  of  General 
Literature,  X).  The  most  recent  paper  is  by  Albert 
Maassen  (Arb.  a.  d.  Kais.  Gesundh.,  Bd.  XXI,  H.  3, 


Fig.  18* 


1904,  p.  377,  6  pi.).  He  found  chloride  of  lithium  specially 
advantageous  for  provoking  these  growths,  which  are  re- 
garded as  teratological.  He  obtained  them  in  24  hoxirs. 

GENERAL   COMMENT. 

Great  care  should  be  paid  to  the  minute  morphology 
of  each  organism,  not  only  in  the  host-plant  but  also  in 
a  variety  of  cultures,  old  and  young,  so  that  a  body  of 
knowledge  more  exact  than  we  now  possess  shall  be  grad- 
ually accumulated  for  differential  and  systematic  purposes. 
Careful  drawings  and  photographs  should  be  made.  The 
Abbe  camera  is  a  great  help  in  making  drawings  (fig.  121). 
For  such  study  the  Zeiss  apochromatic  lenses  and  com- 
pensating oculars  can  not  be  recommended  too  highly, 
particularly  the  16  mm.,  with  the  12  and  18  compensating 
oculars  for  studying  the  margins  of  colonies,  and  the  2  mm. 
1.30  n.  ap.,  with  the  8  and  12  compensating  oculars  for  the 
more  detailed  study  of  the  individual  rods.  The  writer  has 
also  made  much  use  of  the  Zeiss  3  mm.  1.40  n.  ap.  apochro- 
matic objective.  The  Zeiss  screw,  or  filar,  micrometer  com- 
bined with  a  No.  12  compensating  ocular  (fig.  23)  will  be 
found  very  useful.  For  photographic  purposes  the  projec- 
tion oculars  or  the  4  or  6  compensating  oculars  may  be  used. 
Robert  Koch  was  entirely  correct  in  saying :  "A  general 
use  of  photography  in  microscopic  works  would  certainly 
have  prevented  a  great  number  of  unripe  publications." 


Fig.  19.t 


*Fic.  18. — Bacterium  camfestre.  Cover^glass  (smear)  preparation  from  the  vessels  of  a  cab- 
bage plant  received  from  Racine,  Wis.,  Sept.  19,  1896.  Stained  with  carbol-fuchsin.  Drawn  from 
a  photomicrograph.  X  1,000  circa. 

tFic.  19. — Bacterium  campestre  from  an  old  culture  on  23  per  cent  grape-sugar  agar,  showing 
long  filaments.  Cover  stained  I  hour  and  20  minu  tes  in  gentian  violet  ( I  part  saturated  alcoholic 
solution  plus  I  part  water).  Many  of  the  rods  stained  feebly.  Tube  inoculated  June  30,  1898. 
Cover  prepared  Aug.  8.  Drawn  directily  from  the  slide.  X  1,000. 


24  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

Good  photomicrographs  should  be  seciired  if  possible.  Koch's  first  photo- 
micrographs were  of  various  enlargements.  He  afterwards  recommended  X  1,000 
as  the  standard  magnification,  but  X  1,500  and  X  2,000  are  also  convenient  sizes  and 
occasionally  X  500  is  better  than  X  1,000.  Most  important  is  it  that  the  exact  mag- 
nification should  always  be  indicated.  The  Zeiss  apochromatic  objectives  are  much 
better  for  photographic  work  than  the  achromatic  ones.  For  very  small  magnifica- 
tions the  writer  has  found  the  old  Zeiss  35  mm.  and  70  mm.  very  useful.  For  the 
same  purpose  the  newer  Zeiss  planars,  series  la  Nos.  1—5  (fig.  122)  are  admirable. 
These  have  sharp  definition  and  a  very  flat  field,  but  not  much  depth  of  focus.  With 
them  objects  several  centimeters  in  diameter  may  be  satisfactorily  photographed  with 
magnifications  from  2  or  3  diameters  to  50  or  more.  The  writer  obtains  as  sharp  a 
focus  as  possible  with  wide-open  diaphragm  and  then  stops  down  about  two-thirds. 


Fig.  20  * 

One  of  the  best  simple  photomicrographic  outfits  is  the  Zeiss  upright  camera 
(fig.  24).  All  apparatus  is  to  be  rejected  which  requires  the  microscope  to  rest  on 
the  same  platform  as  the  camera.  It  should  rest  on  the  table  independent  ot 

the  camera,  unless  a  weak  light  is  used  and  the  exposures 
r\  are  very  long,  in  which  case  a  slight  jarring  is  of  no  great 

(1     consequence.      Direct    sunlight    is    the   best    light,    but 

J}   \i    0    ^~~    A  8      t^ie  ^&ht  °f  tne  °Pen  sky  may  be  used  (with  full  open 
"          <?      9    (J   0      diaphragm)  if  one  is  willing  to  make  5  to  20  minute 
(j     C\         exposures.     Electric  light  is  often  used  by  those  who  live 
\l         in  cloudy  regions  or  who  occupy  rooms  not  exposed  to 
F.   2|  .  the  sun,  but  the  writer  has  had  no  experience  with  it. 

Very  good  pictures  also  may  be  made  by  gaslight  if  the 

Welsbach  burner  is  used.  Ordinary  lamp  light  (kerosene)  is  too  yellow  and  not 
sufficiently  intense.  Photographs  can  be  made 
with  a  kerosene  light,  but  the  time  and  trouble 
involved  make  it  scarcely  worth  while  to 
consider  this  source  of  light.  The  writer  has 
obtained  the  best  results  by  using  direct  sun- 
light  and  slow  isochromatic  plates  behind  Zett- 
now's  light  filter.  Of  course,  with  upright 
cameras  a  dry  light-filter  must  be  used,  such  as 
the  yellow  one  devised  by  Carbutt  or  by  Ives. 
In  using  a  horizontal  apparatus,  such  as  that 
shown  in  plate  5,  the  sine  qua  non  is  to  get  it  properly  leveled  up  and  to  keep  it  so. 

*Fic.  20. — Hollow-ground  slide  with  cover-glass  bearing  hanging  drop  for  examination  under 
the  microscope. 

fFic.  21. — Involution  forms  of  Bacillus  tracheiphilus  from  extremely  ropy  potato  broth.  Drawn 
free  hand ,  X  1,000  circa.  Many  as  large  as  8  by  2  micra  and  others  larger.  Nov.,  1894. 

JPiG.  22.—  Y  -shaped  (dichotomously  branched)  bodies  from  the  root-tubercles  of  clover  (Tri- 
folium).  From  a  photomicrograph  by  tfhe  author,  made  from  a  slide  furnished  by  Dr.  Geo.  T.  Moore. 
X  1,500. 


INSPECTION    OF    COLONIES. 


For  the  inspection  of  colonies  and  of  subcultures  in  tubes  the  best  hand-lens 
known  to  the  writer  is  the  Zeiss  aplanat  magnifying  six  times  (fig.  25).  That  magni- 
fying 10  times  is  also  very  useful,  but  will  not  reach  to  the  center  of  an  ordinary  test 
tube.  Those  in  apple-tree  wood  cases  are  in  some  respects  more  convenient  than 
those  provided  with  metallic  swing  covers  (fig.  26). 

The  best  general  work  to  consult  on  the  morphology  of  the  bacteria  is 
undoubtedly  Migula's  System  (see  Bibliog.,  III). 

PHYSIOLOGY. 

In  the  description  of  bacteria  we  are  compelled  to  make  large  use  of  physiolog- 
ical peculiarities,  owing  to  their  very  simple  and  monotonous  morphology.  Within 
the  limits  of  the  genera  now  recognized  the  form  differences  are  so  very  slight 


Fig.  23* 

that  many  bacteria,  e.  g.,  Bacillus  coli,  B.  cloaca,  B.  suipestifer,  B.  typhosus, 
B.  amylavorus,  etc.,  are  indistinguishable  under  the  microscope.  In  mixed  cultures, 
or  stained  preparations,  no  one  could  distinguish  one  from  the  other  with  any  cer- 
tainty, and  in  pure  cultures  of  unknown  origin  certain  identification  by  means  of 
the  microscope  would  be  equally  impossible.  Nevertheless,  these  same  forms  are  so 
widely  different  in  their  behavior  in  culture  media,  in  their  pathogenic  properties,  in 
their  relation  to  heat,  air,  antiseptics,  etc.,  that  we  are  certainly  warranted  in  regard- 
ing them  as  distinct  species,  using  the  word  "  species  "  in  its  common  acceptation. 
These  well-ascertained  facts  should  not,  however,  lead  one  to  neglect  slight  differ- 
ences of  form,  even  when  they  can  be  expressed  only  in  fractions  of  a  micron.  On 


*Fic.  23. — Zeiss  compensating  ocular  No.  12  with  screw-filar  micrometer. 


26 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


the  contrary,  as  much  as  possible  should  be  made  out  of  morphology,  particularly 
that  of  the  living  organism,  and  in  this  connection  the  recent  efforts  of  Migula  and 
Fischer  are  especially  deserving  of  commendation. 

MOTILITY. 

If  motile,  determine  kind  of  motion  and  rapidity 
(margin  of  small  hanging  drops  on  thin  covers  sus- 
pended over  hollow-ground  slides).!  The  cover  may 
be  prevented  from  sliding  by  touching  one  edge  with 
a  very  little  vaseline  or  cedar  oil ;  if  too  much  is 
used  it  runs  under,  mixes  with  the  hanging  drop,  and 
spoils  the  mount,  and  possibly  in  the  end  the  objec- 
tive is  ruined,  if  the  student  continues  to  search  for  a 
clear  field.  The  beginner  is  very  apt  to  mistake 
Brownian  movement  for  self-motility.  It  sometimes 
requires  very  careful  observation  to  be  quite  certain. 
Rods  which  appear  to  be  motionless  will  sometimes 
be  seen  to  dart  away  quite  suddenly  if  watched.  In 
some  species  young  cultures  are  much  more  apt  to 
be  motile  than  old  ones  ;  in  others  motility  appears 
to  be  an  almost  constant  characteristic.  The  move- 
ments of  bacteria  are  sometimes  quite  characteristic 
for  particular  sorts.  They  may  be  slow  or  rapid 
tumbling  motions  centering  in  the  shorter  axis,  or 
straight  or  sinuous  slow  or  rapid  darting  move- 
ments in  the  direction  of  the  longer  axis,  with 
rotation  on  this  axis.  The  media  of  Hiss  ('97,  Bib- 
Hog.,  XVI)  and  of  Stoddart  ('97,  Bibliog.,  XVI)  are 
sometimes  useful  for  distinguishing  macroscopically 
between  motile  and  non-motile  forms.  The  former 
spread  as  a  thin  layer  over  the  whole  surface,  the 
latter  pile  up  in  restricted  areas  around  the  points 
of  inoculation.  The  student  should  not  remain  con- 
tent with  merely  determining  motility,  but  when  this 
has  been  settled  he  should  turn  his  attention  to 
staining  the  organs  of  motion. 


Fig.  24* 


*Fic.  24— Upright  Zeiss  camera  for  photomierographic  work.  The  cup  (a)  slips  over  the  end 
of  the  microscope  and  forms  a  light-tight  connection  with  the  bellows  without  touching  it.  The 
microscope  rests  on  the  table  independent  of  the  camera.  The  stout  rod  turns  freely  in  the  socket 
X  and  is  locked  in  place  by  a  set-screw  on  the  side  opposite  the  observer.  The  height  is  about 
45  inches. 

tLehmann  and  Fried  (Arch.  f.  Hyg.,  Bd.  XL VI,  1903,  p.  3")  found  the  swiftest  movement  of 
bacteria  to  be  I  mm.  in  22  seconds ;  the  slowest  I  mm.  in  222  seconds ;  average :  cholera,  I  mm.  in 
3414  seconds;  typhoid,  I  mm.  in  56  seconds;  B.  vulgare,  I  mm.  in  73  seconds;  B.  subtilis,  I  mm.  in 
40  seconds ;  B.  megaterium,  I  mm.  in  2  minutes  1 1  seconds. 


PLATE  5. 


Large  horizontal  Zeiss  photomicrographic  outfit  ready  for  use, 

except  that  when  photographing  the  curtain  n  raised  and  the  mirror  is  placed  farther  away,  i.  e..  out  of  the  south  window  on  the  triangular 
extension  shown  on  the  front  table  at  the  right.  In  the  newer  forms  each  table  top  may  be  railed  oe  lowered  at  will.  There  is  aUo  a 
device  for  raising  or  lowering  the  plate  on  which  the  microscope  rests. 


STUDY   OF   COLONIES. 


GROWTH. 

The  manner  of  growth  and  rapidity  of  growth  at  given  temperatures  in  hanging 
drops  and  also  on  the  margin  of  young  colonies  on  plates  of  nutrient  gelatin  and 

agar  of  varying  density  should  be  determined. 
Frequently  characteristic  and  interesting  ar- 
rangements of  the  rods  forming  the  surface 
layers  of  the  colony,  especially  when  it  is 
young,  may  be  discovered  by  means  of  a 
direct  inspection  of  the  colonies  under  low 
powers  of  the  microscope  or  by  means  of 
cover-glass  impressions.  Covers  are  carefully 
placed  on  the  colony,  removed,  dried,  flamed, 
and  stained.  There  are  also  often  curious 
Fig.  25*  arrangements  of  the  deeper  layers  of  the 

surface  colony.  In  direct  examination  the  colonies 
should  be  viewed  by  reflected  as  well  as  by  trans- 
mitted light.  Drawings  or  photographs  of  surface 
colonies  should  be  made  under  low  or  medium 
magnifications.  By  a  little  practice  using  Lister's 
dilution  method  ('78,  Bibliog.,  XVII),  hanging- 
drops  containing  a  single  bacterium  for  study  under 
the  microscope  may  be  obtained  with  Beyerinck's 
capillary  drop-flask  ('91,  Bibliog.,  XVII). 

CHEMOTROPISM. 


Fig.  26.  t 


On  the  general  subject  of  chemotropism,  see  papers  by  Pfeffer,  Miyoshi, 
Jennings,  Buller,  Rothert,  etc.  Jennings  maintains  that  contact  irritation  inducing 
motor  reflex  is  responsible  for  movements  which  were  formerly  attributed  to  chemical 
stimulus.  Consult  Jennings,  "  Contributions  to  the  study  of  the  behavior  of  lower 
organisms,"  Carnegie  Institution  of  Washington,  1904,  and  especially  Jennings  and 
Crosby,  "The  manner  in  which  bacteria  react  to  stimuli,  especially  to  chemical 
stimuli,"  Am.  Jour.  Physiol.,  Vol.  VL,  pp.  31-37,  and  Jour.  Roy.  Mic.  Soc.,  1902, 
p.  88.  Spirillum  volutans  was  used  in  the  tests. 

REACTION   TO  STAINS. 

Proper  staining  is  a  very  important  part  of  the  study  of  bacteria.  Its  founda- 
tion principle  is  the  fact  that  the  bacteria,  in  a  living  vegetative  condition,  all  show 
a  great  affinity  for  the  basic  anilin  dyes.  Spores  ordinarily  show  no  such  affinity, 
but  may  be  made  to  take  up  stains  by  acting  on  them  with  strong  acids  or  alkalis, 
or  by  heating  them  very  hot.  Flagella  also  show  no  affinity  for  stains  until  acted 

*Fic.  25. — Hand  Jens  suitable  for  examining  bacterial  cultures.  Zeiss  aplanat  magnifying  six 
times.  Tiliree-'four.ths  natural  size. 

tFig.  26. — Zeiss  swing-cover  aplanat  magnifying  six  times.  This  is  now  sent  out  in  a  neat  little 
chamois-skin  purse.  About  two-thirds  natural  size. 


28 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


on  by  severe  reagents,  when  they  may  be  stained  in  mordanted  solutions  or  in  dyes 
which  have  been  preceded  by  a  mordant.  The  outer  wall  of  the  bacterium  generally 
reacts  to  stains  in  the  same  way  as  the  flagella,  i.  e.,  it  usually  remains  unstained. 

Staining  media  may  be  roughly  divided  into  four  groups :  (a)  Simple  stains 
dissolved  in  water,  e.  g.,  fuchsin  (basic),  gentian  violet,  methylene  blue  ;  (b)  alcoholic 
solutions  and  various  complex  stains,  e.  g.,  saturated  alcoholic  solutions  of  anilin 
dyes,  alcohol-iodine,  iodine  potassium  iodide,  Russow's  cellulose  test,  Ziehl's  carbol- 
fuchsin,  Loemer's  alkaline  methylene  blue,  Ehrlich's  anilin-water  gentian  violet, 
Gabbett's  stain,  Gram's  method,  Delafield's  hsematoxylin,  Ehrlich's  acid  heema- 
toxylin,  Heidenhain's  iron-hsematoxylin,  Fleming's  triple  stain;  (c)  flagella  and 
capsule  stains,  e.  g.,  Loemer's  stain,  Moore's  modification,  Fischer's  modification, 
Bunge's  stain,  L,6wit's  stain,  van  Ertnengem's  nitrate  of  silver  method,  Zettnow's 
gold  method,  etc.;  (d)  stains  for  spores,  e.  g.,  prolonged  exposure  to  simple  stains 

dissolved  in  water  (which  should 
have  little  effect),  steaming  carbol- 
fuchsin  with  methylene  blue  for 
contrast,  etc.  (see  "  Formulae  "  and 
Bibliography  of  General  Literature 
under  "Flagella,"  "Spores,"  etc.,  for 
various  observations  on  staining). 

Griibler's  stains  are  preferred. 
Cover-glasses  should  be  clean  and 
free  from  fat,  traces  of  which  are 
easily  removed  in  a  Bunsen  flame. 
A  little  experience  is  necessary  in 
flaming  thin  covers  in  order  not  to 
crack  them.  They  may  be  passed 
through  the  flame  three  times,  wait- 
ing a  moment  or  two  after  each  pass 
for  them  to  cool.  The  student  should 
see  that  the  water  used  in  making 
Fig. 27*  the  cover-glass  preparations  or  the 

stains  does  not  itself  contain  bacteria.  It  is  usually  wise  first  to  dry  a  drop  of 
the  water  on  the  cover  and  stain  without  addition  of  the  bacteria.  Eternal  vigi- 
lance is  the  price  of  trustworthy  results.  It  is  best  to  make  all  mounts  on  cover- 
glasses  of  a  known  and  uniform  thickness  (0.15  mm.).  Many  a  good  preparation  has 
been  spoiled  for  examination  with  lenses  of  a  short-working  distance  by  mounting 
under  a  thick  cover-glass,  and  sometimes  the  lens  itself  has  been  ruined  in  the 
attempt  to  focus.  The  thickness  of  covers  often  varies  greatly  from  the  statements 
of  dealers,  and  they  should  not  be  accepted  until  tested  with  a  reliable  cover-glass 
measurer  (fig.  27). 


*PiG.  27.- — Zeiss  cover-glass  -measurer.    The  cover  in  place  shows  a  registered  thickness  of  0.18 
mm.    Fractions  of  an  dnch  are  also  registered  on  fhis  instrument. 


REACTION    TO    STAINS.  29 

To  determine  whether  bacteria  are  properly  stained  examine  with  the  diaphragm 
of  the  condenser  wide  open.  If  they  can  not  be  seen  distinctly  with  this  flood  of 
light  they  are  not  well  stained.  The  bacteria  should  be  well  separated  on  the  cover 
and  deeply  stained,  while  the  background  should  be  very  free  from  stain. 

Dr.  Weigert  seems  to  have  been  the  first  to  use  anilin  stains  for  the  demon- 
stration of  bacteria  in  tissues.  This  was  about  1875.  Since  that  time  staining  in 
tissues  has  been  worked  up  carefully  for  bacteria  causing  animal  diseases,  but  very 
little  is  known  respecting  best  methods  of  staining  bacteria  in  vegetable  tissues. 
The  difficulty  lies  in  the  fact  that  the  tissues  of  the  higher  plants  often  take  the 
basic  anilin  stains  as  readily  as  the  .bacteria  and  retain  them  even  more  tenaciously. 
Special  remarks  may  be  looked  for  under  particular  diseases. 

CULTURE   MEDIA. 
NUTRIENT  GELATIN. 

(«)  Plate  Ctiltures. — Colonies,  young  and  old,  buried  and  superficial,  crowded 
and  wide  apart,  should  be  examined  for  color,  translucency  or  opaqueness,  shape, 
thickness  of  the  surface  growth,  and  character  of  the  margin.  They  should  also 
be  studied  under  low  powers  of  the  compound  microscope  for  lobes,  branches, 
granulations,  wrinkles,  flecks,  concentric  rings,  radial  filaments,  arrangement  of  the 
dividing  rods  on  the  margin  of  the  colony,  iridescence,  etc.  The  microscopic 
appearance  of  the  surface  colony  during  the  first  48  hours  is  often  different  from 
that  later  on.  The  rapidity  of  growth  should  be  compared  with  that  of  some 
common  and  easily  accessible  organism,  e.g.,  Bacillus  coli,  B.  amylovorus,  Bacterium 
campestre.  The  comparative  rate  of  growth  of  buried  and  surface  colonies  should 
also  be  carefully  noted.  How  is  the  appearance  of  the  colony  changed  by  increasing 
the  amount  of  gelatin,  or  varying  the  brand  of  gelatin?  Are  the  surface  colonies 
viscid,  or  can  they  be  lifted  bodily  in  one  mass  from  the  substratum  ? 

(&)  Stabs. — The  nature  of  the  surface  growths  and  of  the  deeper  growths  should 
be  carefully  examined.  Is  there  any  marked  tendency  of  the  latter  to  grow  down- 
ward or  outward  into  the  body  of  the  gelatin,  either  in  distinct  masses  or  as  a  dif- 
fused cloudiness?  Observe  effect,  if  any,  on  growth  when  the  gelatin  is  acid  or  only 
feebly  (litmus)  alkaline.  If  liquefaction  of  the  gelatin  occurs,  note  its  rapidity  and 
whether  it  is  mostly  restricted  to  the  surface  or  is  equally  rapid  along  the  line  of 
the  stab  in  the  depths ;  note  also  whether  the  liquefied  gelatin  is  clear  or  cloudy  in 
tubes  which  have  not  been  shaken,  and  whether  a  pellicle  has  formed  on  its  surface. 
Liquefaction  may  be  very  rapid  (taking  place  within  a  few  hoiirs),  may  occur  after 
three  or  four  days,  may  be  long-delayed  and  feeble  (only  visible  after  some  weeks), 
or  may  not  occur  at  all.  It  is  the  cases  of  feeble  and  long-delayed  liquefaction 
which  lead  to  contradictory  statements  on  the  part  of  different  observers,  and  con- 
sequently cultures  should  remain  under  observation  for  a  considerable  time  and  on 
a  variety  of  gelatins.  Various  substances  interfere  with  liquefaction.  Determine 
whether  liquefaction  can  be  prevented  by  the  addition  of  grape-sugar  or  cane-sugar 
(10  per  cent).  Look  for  gas-bubbles,  for  crystals,  for  any  fluorescence  or  staining 
of  the  medium  (green,  brown).  Inasmuch  as  the  growth  of  some  bacterial  plant 


BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 


parasites  is  restrained  by  some  nutrient  gelatins  which  are  neutral  or  only  feebly 
alkaline  to  litmus,  it  is  advisable  to  add  to  a  part  of  the  stock  more  caustic  soda  than 
is  commonly  used,  z.  <?.,  enough  to  render  it  neutral  to  phenolphthalein  (strongly 
alkaline  to  neutral  litmus),  especially  if  gelatin  is  selected  as  the  first  medium  for 
isolation  experiments  ;  otherwise  perplexing  failures  may  result. 

(c)  Streaks. — Record  the  character  of  the  streak,  whether  wet  or  dry,  smooth, 
wrinkled,  or  rough,  thin  or  piled  up,  margin  well  defined  or  indistinct.  Note  also 
whether  the  surface  is  ever  iridescent,  whether  growths  are  sent  down  from  the  under 
surface  into  the  substratum,  whether  the  streak  spreads  rapidly  and  widely  over  the 

surface  or  very  slowly.  The  sur- 
face behavior  depends  to  some 
extent  on  the  motility  of  the 
organism,  on  the  amount  of  water 
in  the  surface  layers,  z.  e.,  whether 
the  slants  are  fresh  or  old,  and  on 
the  amount  of  gelatin  in  the  me- 
dium, which  in  temperate  climates 
should  usually  be  loper  cent,  but 
may  be  15  or  even  20  per  cent. 
By  minimizing  heat  in  prepara- 
tion and  by  increasing  the  quan- 
tity of  gelatin  to  20  or  30  per 
cent  a  medium  may  be  obtained 
which  will  remain  solid  at  30°  C. 
Growth  is  less  satisfactory,  how- 
ever, on  such  a  dense  medium, 
or  at  least  was  in  the  few  tests 
made  by  the  writer.  Chester  has 
applied  the  ordinary  botanical 
terminology  to  the  varying  mar- 
gins of  colonies,  etc.,  and  has  pub- 
lished some  useful  figures  ('or, 
Bibliog.,  III). 

No  siibstance  used  in  the  bac- 
F>g-  28.*  teriological  laboratory  is  so  uncer- 

tain and  variable  in  its  composition  as  gelatin.  The  gelatin  from  different  factories 
varies  greatly  and  hardly  any  two  batches  from  the  same  factory  are  alike.  One  glue 
chemist  has  defined  gelatin  as  "  80  per  cent  glue,  10  per  cent  dirt,  and  10  per  cent 
doubt."  It  varies  greatly  in  its  melting  point  and  power  of  setting,  and  in  amount 
of  peptones  and  albumoses  it  may  contain,  which  is  sometimes  large.  It  always  con- 
tains calcium  salts  and  phosphates,  which  are  often  antiseptic,  and  the  nature  of  which 
varies  according  as  hydrochloric  or  sulphurous  acid  has  been  used  in  its  manufacture. 
Formaldehyde  is  sometimes  added  to  it,  we  are  told  ;  and  occasionally  agar  also,  it  is 

*Fic.  28.— Nelson's  photographic  gelatin  No.  I.    Recommended  for  bacteriological  use. 


VARIABILITY   OF   GELATIN.  3! 

said,  is  added  to  certain  table  gelatins  to  increase  their  body.  Gelatin  also  contains 
a  variety  of  decomposition  products  due  to  the  growth  in  it  of  various  fungi  and 
bacteria  while  it  is  in  the  vats  or  in  the  drying-house.  If  there  is  any  delay  in  the 
drying  it  is  spotted  all  over  with  molds  and  bacteria.  It  also  contains  some  wax  or 
grease,  used  to  anoint  the  surface  on  which  it  is  spread  to  dry,  and  this  wax  or 
grease  is  probably  also  a  variable  substance.  Gelatins  also  polarize,  it  is  said,  in 
many  different  ways.  An  absolutely  pure  gelatin  of  uniform  character  for  bacterio- 
logical purposes  is  not  to  be  had.  That  which  perhaps  comes  the  nearest  to  it  and 
which  is  here  recommended  is  Nelson's  gelatin,  made  in  L,ondon  and  well  known  to 
the  makers  of  photographic  dry-plates,  who  use  it  in  large  quantities.  It  comes  in 
two  grades,  a  hard  and  a  soft,  and  costs  about  $1.25  per  pound.  No.  i,  that 
which  I  like  best,  comes  in  shreds  resembling  "  excelsior  "  used  for  packing  (fig.  28). 
No.  3,  which  comes  in  long,  broad  strips,  contains  much  cell  detritus,  etc.,  and  filters 
with  difficulty.  Other  expensive  gelatins,  said  to  be  of  quite  uniform  quality,  are 


Fig.  29* 

Lichtdruck  gelatin,  made  by  Carl  Creutz,  Michelstadt,  in  Hesse,  and  Geneva  Red 
Cross  gelatin  made  by  Winterthur,  in  Switzerland,  under  direction  of  Dr.  Eder,  of 
the  Imperial  Institute  of  Vienna  (Cockayne ). 

NUTRIENT  AGAR. 

Agar,  or  agar-agar,  as  it  is  usually  called,  from  a  Malay  word  meaning  "  vege- 
table," is  a  manufactured  product  obtained  from  various  sea-weeds  growing  in 
Chinese  and  Japanese  waters.  Various  species  are  used  as  food  and  the  trade  is  con- 
siderable. It  usually  comes  into  the  hands  of  the  bacteriologist  as  long,  slender, 
yellowish-white  strips  (fig.  29)  or  as  blocks  (fig.  30),  or  more  especially  in  recent 
years,  in  the  form  of  a  gray-white  fine  powder  of  European  manufacture  (fig.  33). 
It  is  reputed  to  be  the  product  of  species  of  Gelidium  (figs.  31  and  32). 


*Fic.  29. — The  kind  of  agar-agar  usually  employed  in  bacteriological  work.  This  is  a  manu- 
factured product  known  to  die  Japanese  as  slender  "  Kanten."  The  figure  represents  first  quality 
"  Kanten,"  in  unbroken  package.  (Courtesy  of  Dr.  Hugh  M.  Smith,  Deputy  Commissioner  of  the 
United  States  Bureau  of  Fisheries,  who  brought  the  package  with  him  from  Japan.) 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


Of  the  Japanese  algse  in  this  group  the  following,  according  to  Rein  (pp.  81-82), 


deserve  special  mention : 

(i.)  Chondrus  punctatus  Sur. 

(2.)  Gigartina  tenella  Harvey;  Jap.  Ogo. 

(3.)  G.  intermedia  Sur. 

(4.)  Gloiopeltis  tenax  Kg.  (Sphaerococcus 
tenax  Ag.) 

(5.)  Gl.  capillaris  Sur.;  Jap.  Shiraga-nori. 

(6.)  Gl.  coliformis  Harv. ;  Jap.  Kek'Kai. 

(7)  Gl.  intricata  Sur. ;  Jap.  Fu-nori. 

(8.)  Gelidium  corneum  Lamouroux;  Jap. 
Tokoroten-gusa. 

(9.)  G.  Amansii  Lamour. 


(10.)   G.  cartilagincum   Gail. 

(n.)  G.  rigidum  Grev. ;  Jap.  Tosaka-nori,  i.  e., 
cockscomb  alga;. 

(12.)  Sphaerococcus  confervoidcs  A.;  Jap. 
Shiramo. 

(13.)  Gymnogongrus  ftabclliformis  Harv.; 
Jap.  Home-nori. 

(14.)  G.  japonicus,  Sur.;  Jap.  Tsuno-mata. 

(15.)  Kallymenia  dentata;  Jap.  Tosaka-nori. 

(16.)  Porpliyra  vulgaris  Ag. ;  Jap.  Asakusa- 
nori. 


Fig.  30.* 

Agar-agar  is  a  neutral  or  nearly  neutral  substance  which  is  converted  by  boil- 
ing with  water  into  a  stiff  jelly  that  hardens  in  i  per  cent  solution  at  39°  to  40°  C., 
and  is  not  easily  liquefied  either  by  the  growth  of  organismsf  or  by  heat  less  than 
that  of  boiling  water.  It  is  a  kind  of  vegetable  glue  forming  a  good  matrix  for 
various  nutrient  substances.  A  chemical  analysis  by  Karten  (Descript.  Cat.  Int. 
Health,  Exhib.,  London,  1884)  gave  the  following  proximate  composition  :  11.71  per, 
cent  nitrogenous  matter  (albumen  [?]),  62.05  per  cent  non-nitrogenous  matter  (evi- 
dently glue,  the  pararabin  of  Reichardt),  3.44  per  cent  ashes,  and  22.80  per  cent  water. 

*Fic.  30.— Another  form  of  agar-agar  known  to  the  Japanese  as  square  "  Kanten."  The  bulk  of 
this  goes  to  Holland,  where  it  is  used  for  clarifying  schnapps.  Courtesy  of  Dr.  Hugh  M.  Smith. 
The  actual  slize  of  these  sticks  is  about  10%  by  2^2  by  1^4  inches. 

tMetcalf  has  described  a  bacillus  which  slowly  softens  k,  and  the  writer  has  observed  similar 
phenomena. 


PREPARATION    OF    NUTRIENT   AGAR. 


33 


For  a  full  account  of  Japanese  methods  of  making  agar-agar  consult  a  paper 
entitled  "The  Seaweed  Industries  of  Japan,"  by  Dr.  Hugh  M.  Smith,  in  the  Bulletin 
of  the  United  States  Bureau  of  Fisheries  for  1904. 

In  addition  to  beef  bouillon, 
or  in  place  of  it,  various  sub- 
stances, organic  and  inorganic, 
may  be  added  to^the  agar  with 
advantage.  The  writer  makes 
much  use  of  litmus-lactose  agar, 
which  is  made  out  of  ordinary 
nutrient  agar  by  adding  i  per 
cent  milk-sugar  and  enough 
pure  litmus  water  to  give  a  pur- 
ple-red color.  Glycerin-agar, 
waltose-agar,  etc.,  may  be  made 
up  with  any  amount  of  the  sub- 
stance desired,  generally  i  or  2 
per  cent. 

Formerly  it  was  difficult  to 
filter  agar  perfectly  clear  and  it 
was  therefore  used  less  than 
gelatin,  but  in  recent  years  it 
has  been  discovered  that  this 
difficulty  may  be  overcome  if 
the  agar  is  first  brought  into 
complete  solution  by  prolonged 
boiling  or  by  a  short  boiling  at 
a  temperature  somewhat  above 
100°  C,  e.g.,  uo°C. 

The  writer  formerly  obtained 
filtered  clear  agar  by  soaking 
the  snipped  agar  in  5  per  cent 
acetic-acid  water  for  some  hours, 
after  which  a  thin  cloth  was  tied 
over  the  mouth  of  the  beaker 
securely,  and  tap  water  allowed 
to  run  into  it  for  an  hour  or  more 
/'.  e.,  until  all  trace  of  acid  was 
removed.  The  softened  agar 
was  then  put  into  the  bouillon, 
boiled  for  two  hours,  and  finally 
filtered  through  S.  &  S.  filter 


6 
Fig.  3 1* 


*Fic.  31. — Red  sea-weeds  from  which  agar-agar  is  manufactured,  a,  Gelidium  corncum  Lam., 
one-third  natural  size;  b,  Gelidium  subcostatum  Lam.,  one-half  natural  size.  From  a  colored  Jap- 
anese chart  showing  "  The  principal  aquatic  plants  of  Japan,"  supposed  to  be  an  official  publication. 
Original  in  the  library  of  the  United  States  Fish  Commission. 


34 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


paper,*  using  a  hot-water  funnel.  Later  he  followed  Schutz's  method  ('92,  Bibliog., 
XVI),  which  is  a  very  good  one.  This  consists  in  cutting  the  agar  into  small  bits 
and  first  heating  it  very  hot  in  a  beaker  or  enanieled-iron  dish  in  a  minimum  quantity 
of  water  or  beef-bouillon  over  a  hot  Bunsen  flame  with  constant  and  rapid  stirring  and 


Fig.  32.f 


*The  folded  filter  papers  are  the  most  convenient  (fig.  34).  These  filter  papers  give  the  starch 
reaction  (blue)  with  iodine,  and  reduce  Fehling's  solution  on  being  boiled  in  it. 

tFiig.  32. — -Unnamed  species  of  red  sea-weeds  (GV/jrf»«m)  furnishing  agar-agar.  From  a  Japan- 
ese chart  showing  "  Tlie  principal  aquatic  plants  o  f  Japan,"  supposed  to  be  an  official  publication. 
One-half  natural  size.  Original  in  library  of  United  States  Fish  Commission. 


PREPARATION    OF    NUTRIENT   AGAR. 


35 


*X3fe  • '  '  '  ' 

•..••'•••     .      '       •  * 
•   ..._. ,       — *-•  ~^  ii  n^» 


occasional  additions  of  small  quantities  of  water  until  it  is  thoroughly  cooked  in 
the  form  of  a  thick  mush.  It  is  then  put  into  the  remainder  of  the  water  or  bouil- 
lon and  subjected  to  streaming  steam  for  two  hours,  after  which,  if  the  first  heating 
was  sufficient,  it  niters  readily  without  the  use  of  a  hot-water  filter,  or  the  necessity 
of  keeping  it  in  the  steamer  during  the  filtering.  The  stirring  rod  must  touch  all 
parts  of  the  bottom  of  the  dish  exposed  to  the  flame,  every  few  seconds  during  the 
preliminary'  heating,  otherwise  the  agar  will  burn  on  and  be  spoiled.  On  some 
^s<~^-^  accounts  it  is  best  to  begin 

operations  with  beakers  rather 
than  the  enameled  iron  dishes. 
In  this  way  all  likelihood  of 
using  burned  agar  is  avoided, 
since  the  moment  the  agar 
burns  on  the  beaker  cracks  and 
the  agar  is  spilled.  For  bacte- 
riological use  agar  should  be 
clear,  not  cloudy  or  filled  with 
unremoved  precipitates. 

The  writer  now  employs  an 
autoclave  and  uses  an  agar  flour 
procured  from  Lautenschlager  or 
Merck  (fig-33).  If  one  has  an  au- 
toclave the  preliminary  heating 
of  the  agar  in  an  open  dish  with 
a  minimum  quantity  of  water 
and  all  the  subsequent  stages 
may  be  dispensed  with  and  the 
entire  process  carried  on  in  the 
autoclave,  unless  it  is  known 
or  suspected  that  media  heated 
in  the  autoclave  are  less  well 
adapted  to  the  growth  of  par- 
ticular organisms  than  those  pre- 
pared at  1 00°  C.  The  amount 
of  agar  added  to  the  culture 
fluid  is  usually  i  per  cent.  On 
the  making  of  nutrient  agar 


Fig.  33.* 


consult  "Formula:,"  and  the  various  standard  text-books. 

Is  there  any  difference  in  the  appearance  of  colonies  when  grown  at  5°  to  10°, 
15°  to  20°,  and  30°  to  37°  C.?  Observe  the  amount  of  precipitate  that  collects  in 
the  fluid  in  the  V.  For  other  observations  as  to  growth  on  this  substratum  see 
"Gelatin."  Every  organism  should  be  studied  in  numerous  Petri-dish  poured-plate 


*Fic.  33. — Agar-agar  flour  as  received  from  European    manufacturers, 
agar  flour. 


Package   of   Merck's 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


cultures.  Too  many  plate  cultures  can  scarcely  be  made.  Dishes  with  flat  and  very 
thin  bottoms  (0.3  mm.)  are  desirable  for  some  purposes,  but  are  difficult  to  procure. 
For  quantitative  work,  plates  with  flat  bottoms  are  necessary,  and  when  photographs 
are  likely  to  be  wanted  plates  must  be  selected  which  do  not  have  rings,  wavy  places 
or  other  flaws  in  the  glass  on  the  bottom.  There  is  room  for  much  improvement 
in  the  quality  of  the  Petri  dishes  now  on  the  market. 

The  stxident  is  advised  to  use  agar  media  for  all  general  laboratory  work.  When 
he  has  learned  the  behavior  of  an  organism  on  nutrient  agar,  he  may  then  try  gelatin. 
Do  any  of  the  organisms  under  observation  soften  or  liquefy  the  medium  ? 

Agar  roll  cultures  may  be  made  in 
test  tubes  readily  if  the  amount  of 
fluid  agar  is  reduced  to  one-half  cubic 
centimeter. 

When  colonies  are  to  be  counted, 
special  pains  must  be  taken  to  dis- 
tribute the  gelatin  or  agar  uniformly 
over  the  bottom  of  the  dish. 

Various  persons  —  Pake,  Jeffer, 
Weiss,  Mace",  et  al. — have  devised 
ruled  plates  for  counting  the  number 
of  colonies  of  bacteria  in  Petri-dish 
poured  plates.  The  writer  prefers  to 
count  by  square  centimeters  or  frac- 
tions thereof.  When  the  plate  is  sown 
thin  enough,  the  entire  number  of 
colonies  should  be  counted.  When 
it  is  very  dense,  the  average  may  be 
taken  of  ten  square  centimeters  se- 
lected with  care,  provided  the  bottom 
is  flat,  otherwise  the  whole  plate  must 
be  counted.  If  the  counting  plate 
is  to  be  placed  under  the  dish,  it  may 
be  opaque,  i.  e.,  a  black  surface  with 
white  lines,  not  the  reverse.  If  it  is 
to  be  placed  on  top  of  the  dish,  the  latter  preferably  bottom  up,  then  it  should  be 
of  glass  or  some  other  transparent  substance.  The  spaces  may  then  be  ruled  on 
with  a  diamond,  or  drawn  on  in  very  fine  black  lines  with  India  ink.  The  gelatin 
film  of  an  unexposed,  fixed  photographic  dry-plate  is  a  very  good  surface  for  holding 
the  ink.  For  counting  colonies  on  very  densely  sown  plates,  the  writer  has  found 
convenient  a  rectangle  20  mm.  by  5  mm.  divided  into  tenths. 

SIUCATK  JBW.Y. 

In  recent  years,  in  the  hands  of  Winogradsky  and  his  students,  silicate  jelly  has 
played  an  important  part  in  the  isolation  of  various  organisms,  which  do  not  take 


Fig.  34* 


*FiG.  34— Folded  fiker  papers  made  by  Schleioher  &  SdiiiU. 


PREPARATION    OF   SILICATE   JELLY. 


37 


kindly  to  culture  media  containing  animal  and  vegetable  products.  It  is  desirable 
also  for  exact  experiment  with  other  organisms.  It  may  be  used  in  Petri  dishes  or 
flasks,  or  slanted  in  test  tubes.  Along  with  some  disadvantages,  e.  g.,  tendency  to 
split,  it  has  a  number  of  valuable  characteristics,  not  least  among  which  is  the  fact 
that  it  enables  one  to  offer  the  organism  a  solid  substratum  which  is  at  the  same 
time  purely  synthetic.  It  is  generally  considered  to  be  very  difficult 
to  make,  but  by  following  the  most  recent  directions  of  Ome'lian- 
ski  ('99,  Bibliog.,  XXV),  and  especially  certain  slight  modifications 
introduced  by  Moore  &  Kellerman  and  by  the  writer  and  his  assist- 
ants, it  can  be  prepared  without  difficulty,  and  to  it  may  be  added 
any  mineral  nutrient  substances  desired.  The  writer  makes  it  in  the 
following  way: 

To  each  100  cc.  HC1  (sp.  gr.  1.10°  Beauine")  is  added  drop  by 
drop  100  cc.  sodium  silicate  (sp.  gr.  1.09),  the  mixture  being  stirred 
continually  with  a  glass  rod.  This  is  now  placed  in  a  collodion  sack 
and  dialyzed  for  some  hours  in  running  water.  To  this  is  then 
added  in  concentrated  sterile  form  whatever  synthetic  culture  medium 
is  desired,  after  which  the  jelly  is  put  into  Petri  dishes  or  test  tubes 
and  sterilized  by  heating  for  three  hours  in  the  blood-serum  oven  (fig. 
45)  on  five  consecutive  days  at  90°  C.,  or  by  one  steaming  in  the 
autoclave  for  15  minutes  at  110°  C.  The  thermo-regulator  shown  in 
ng-  35  is  useful  for  maintaining  a  constant  high  temperature  in  the 
oven.  The  oven  must  also  contain  some  water  in  a  capsule  or  beaker. 
It  is  believed  that  a  more  detailed  account  of  the  manipula- 
tions connected  with  the  preparation  of  silicate  jelly  will  be  welcome 
to  many.  First  of  all,  one  must  have  dialyzing  sacks.  Collodion 
sacks  are  much  more  convenient  than  parchment  sacks,  since  they 
can  be  prepared  at  any  time,  and  dialysis  takes  place  through  them 
with  great  rapidity.  They  are  useful  for  so  many  purposes  that 
material  for  making  them  should  be  on  hand  in  every  laboratory. 

The  writer  follows  Kellerman  in  making  his  sacks  inside  of  test 
tubes.  These  may  be  large  or  small  according  to  what  the  sacks  are 
to  be  used  for.  If  for  dialyzing  silicate  jelly  in  some  quantity,  it  is 
very  convenient  to  make  the  sacks  inside  of  test  tubes  7  inches  long 
and  having  an  internal  diameter  of  i  inch.  The  first  thing  is  to 
prepare  the  collodion  mixture.  This  is  made  by  dissolving  soluble 
guncotton,  such  as  is  used  by  photographers,  in  a  mixture  of  abso- 
lute alcohol  and  sulphuric  ether.  The  writer  uses  equal  parts  of 
these  two  fluids.  If  too  much  alcohol  is  used,  the  sacks  dry  slowly, 
and  if  too  much  ether  they  are  said  to  become  brittle.  After  some 


Fig.  35.* 


*FiG.  35.— Tollen's  thenmo-regulaitor  for  maintaining  blood-serum  oven  at  80°  to  90°  C.  The 
stem  and  bottom  of  the  bulb  contain  mercury.  The  remainder  of  the  bulb  is  filled  with  glycerin. 
In  the  similar  thermo-regulator  used  for  the  paraffin-ibath  chloroform  replaces  the  glycerin. 
Actual  height,  12  inches.  Chloroform  and  glycerin  are  very  useful  in  such  thermo-regulators  be- 
cause their  coefficient  of  expansion  is  much  greater  than  that  of  mercury.  Toluene  may  also  be 
used  witfi  mercury. 


38  BACTERIA   IN    RELATION    TO   PLANT   DISEASES. 

experimenting  it  was  found  that  5  grams  of  the  clean,  white  guncotton  per  100 
cc.  of  the  fluid  gave  a  solution  very  satisfactory  to  work  with.  About  24  hours  is 
required  to  dissolve  the  guncotton  into  a  homogeneous  mixture,  of  which  there 
should  be  at  least  800  cc.  This  should  be  stored  in  a  cork-stoppered  bottle  of  shape 
convenient  to  hold  in  one  hand.  It  is  then  ready  for  use.  The  clean  test  tube, 
thoroughly  dry  on  the  inside,  is  now  held  in  one  hand  in  a  slanting  position,  mouth 
tip,  while  with  the  other  the  collodion  is  poured  slowly  and  steadily  into  the  tube, 
while  the  latter  is  slowly  rotated.  In  this  way  air-bubbles  are  avoided  and  the  entire 
interior  of  the  tube  is  moistened.  When  this  has  taken  place  and  about  an  inch  of 
fluid  has  accumulated  in  the  bottom  of  the  tube,  the  excess  is  poured  back  into  the 
bottle,  slowly  rotating  the  slanted  tube,  as  before,  so  as  to  cover  again  the  entire 
interior  with  as  imiform  a  layer  as  possible.  When  the  bulk  has  been  poured  back, 
the  tube  is  stood  upright,  mouth  down,  to  drain  on  a  sheet  of  clean  paper.  In  two 
or  three  minutes  it  will  have  drained  sufficiently,  the  excess  of  accumulations  about 
the  mouth  being  wiped  off  on  the  paper  now  and  then.  The  tube  is  then  seized 
and  rotated  in  a  horizontal  position  for  four  or  five  minutes  with  the  mouth  in  the 
draft  of  an  electric  fan,  or  the  rotation  may  be  somewhat  longer  if  no  air-current 
is  available.  A  little  experience  will  tell  when  the  sack  is  dry  enough  to  remove 
from  the  tube.  The  strong  smell  of  ether  must  have  somewhat  subsided  and  the 
collodion  must  not  feel  wet  around  the  mouth  of  the  tube,  as  will  be  the  case  if  the 
layer  of  collodion  is  too  thick  in  places.  If  it  is  taken  out  in  this  condition,  the 
thick,  wet  places  will  become  clouded.  The  collodion  is  now  cut  free  at  the  lips  of 
the  test-tube  by  means  of  a  pin-point  or  other  sharp  instrument  and  the  tube  is  filled 
with  cool  water,  taking  care  to  let  it  also  flow  between  sack  and  wall  of  tube  if  there 
is  any  shrinkage.  In  a  minute  or  two,  if  the  work  has  been  well  done,  the  sack,  free 
from  air-bubbles  and  filled  with  water,  may  be  readily  lifted  out  of  the  tube.  It  is 
then  placed  in  a  jar  of  water,  where  it  remains  until  it  is  ready  to  receive  the  sub- 
stance to  be  dialyzed.  These  sacks  are  quite  tough,  and  there  is  little  danger  of 
tearing  them  during  filling  and  tying. 

When  the  silicate  jelly  or  other  substance  has  been  placed  in  them,  the  mouth 
is  brought  together  and  tied  by  means  of  a  small  rubber  band,  the  elasticity  of  which 
keeps  the  sacks  perfectly  tight.  Silicate  jelly  should  be  dialyzed  for  at  least  1 2  hours, 
and  sometimes  for  24  hours,  if  every  trace  of  salt  must  be  removed.  The  writer 
fills  the  sacks  with  the  silicate  jelly  in  the  afternoon  and  leaves  them  in  running  tap 
water  over  night.  The  next  morning  they  are  taken  out,  their  contents  emptied 
into  a  clean  beaker,  the  nutrient  salts  added,  and  the  fluid  immediately  pipetted  into 
tubes,  flasks,  etc.,  and  sterilized  by  heat.  The  nutrient  substances  should  be  dis- 
solved in  advance,  so  as  not  to  delay  the  preparation  of  the  medium.  They  should 
be  added  for  this  purpose  to  a  minimum  quantity  of  water.  Some  dissolve  slowly, 
and  there  is  a  preferable  order  of  solution,  the  glycerin  being  added  last  in  case  of 
Fermi's  solution. 

For  the  preparation  of  silicate  jelly  a  Beauine"  hydrometer  for  liquids  heavier 
than  water  is  used.  C.P.  hydrochloric  acid  of  any  specific  gravity  is  diluted  with 
distilled  water  until  it  tests  1.10°  on  the  scale  of  the  hydrometer  when  cooled 


PREPARATION    OF    SILICATE   JELLY.  39 

to  60°  F.  Clear  homogeneous  sodium  silicate  of  any  specific  gravity  is  then  mixed 
with  distilled  water  until  it  is  of  sp.  gr.  1.09°  Beaum£  at  60°  F.  A  great  deal  of  water 
must  usually  be  added  to  the  sodium  silicate,  and  the  first  dilution  is  tedious.  For 
example,  100  cc.  of  a  sodium  silicate  of  sp.  gr.  1.42°  required  the  addition  of  750  cc. 
of  distilled  water  to  give  a  fluid  registering  1.07  Beaume".  On  adding  the  fluid 
containing  the  nutrient  salts,  and  hardening,  sodium  silicate  of  sp.  gr.  1.07°  Beaume 
gave  a  rather  too  fluid  medium,  and  sodium  silicate  of  much  higher  sp.  gr.  than 
1.09°  Beaumd  is  apt  to  set  before  it  has  properly  dialyzed,  or  after  adding  the 
nutrient  salts  and  before  it  can  be  tubed  and  slanted.  Several  liters  of  the  diluted 
acid  and  sodium  silicate  may  be  conveniently  made  up  at  one  time.  When  these 
are  ready,  equal  volumes  of  the  two  are  mixed.  This  is  done  by  adding  the 
sodium  silicate  drop  by  drop  to  the  acid,  rather  rapidly,  stirring  meanwhile  with  a 
glass  rod.  The  top  part  of  the  apparatus  shown  in  fig.  146  may  be  used  for  this 
purpose.  The  salty,  acid  fluid  is  now  ready  to  be  placed  in  the  collodion  sacks  for 
dialyzing  in  running  water.  It  is  ready  for  removal  from  the  water  when  it  is  no 
longer  acid  to  litmus  and  shows  only  traces  of  sodium  chloride  remaining.  An 
exposure  to  the  running  water  for  6  hours  is  scarcely  sufficient,  unless  the  sacks  are 
small. 

For  many  purposes  Fermi's  solution  is  a  good  one  to  add  to  the  dialyzed  jelly. 
This  is  made  as  follows,  for  this  purpose:  Freshly-boiled  distilled  water,  100; 
magnesium  sulphate,  o. 2  ;  monopotassium  phosphate,  i.o;  ammonium  phosphate, 
10.0.  Dissolve.  Then  add  glycerin,  45.0. 

The  dialyzed  silicate  jelly  is  now  poured  out  of  the  collodion  sacks  into  a  clean 
beaker  and  brought  to  a  boil  for  a  minute  or  two  over  an  open  flame  (to  drive  off 
the  absorbed  air).  It  is  now  cooled  down  to  50°  C.  and  the  Fermi  added.  If  this 
has  been  dissolved  over  night  it  must  also  be  brought  to  a  boil  and  cooled,  or  have 
the  air  removed  under  an  air-pump  before  adding  it  to  the  silicate  jelly.  To  500  cc. 
of  the  dialyzed  fluid,  90  cc.  of  the  Fermi  may  be  added.  This  is  stirred  with  a  clean 
glass  rod  and  then  quickly  pipetted  into  test  tubes. 

It  is  now  placed  in  the  autoclave  without  delay  in  the  position  desired  and 
heated  for  15  minutes  at  1 10°  C.  To  avoid  tearing  the  surface  of  the  jelly  by  steam, 
the  autoclave  must  be  carefully  shut  steam-tight  as  soon  as  the  air  is  driven  out,  and 
it  must  not  be  opened  until  the  temperature  has  again  fallen  to  100°  C.  It  is  also 
necessary  to  keep  the  autoclave  closed  on  account  of  loss  of  ammonia  from  the 
ammonium  salt.  For  this  reason  it  is  desirable  to  dissolve  the  Fermi  in  freshly- 
boiled  water  and  to  pump  out  any  absorbed  air  rather  than  to  boil  it  out. 

Other  nutrient  salts  may  be  added — Uschinsky's  solution,  etc.  The  writer  has 
had  very  good  success  with  Fermi  for  differential  purposes.  Many  organisms  grow 
remarkably  well  on  this  substratum,  while  others  do  not  vegetate,  or  make  only  a 
scanty  growth. 

The  observations  on  this  medium  are  the  same  as  for  gelatin  or  agar.  Observe 
character  of  growth,  staining  of  substratum  (green,  pink),  etc. 

SOLID  VEGETABLE  SUBSTANCES. 

These  should  consist  of  slant  cylinders  in  cotton-plugged  test  tubes  half  covered 
with  distilled  water  and  steamed  20  minutes  at  100°  C.  on  each  of  three  consecutive 


4O  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

days.  The  addition  of  considerable  water  enables  one  to  keep  the  culture  under 
observation  for  several  months  without  danger  from  drying  out  if  the  cotton  plugs 
are  properly  made.  Drier  culture  media  may  also  be  used.  If  one  wishes  to  do  so, 
the  potato  or  other  substance  may  be  lifted  entirely  out  of  the  water  by  making  a 
constriction  in  the  lower  part  of  the  test  tube,  a  la  Roux,  or  by  thrusting  a  wad  of 
absorbent  cotton  into  the  bottom  of  the  test  tube  before  the  potato  is  introduced. 
The  writer  has  not  found  these  methods  necessary.  In  general,  I  prefer  vegetable 
media  which  have  been  sterilized  in  the  steamer  rather  than  in  the  autoclave. 
The  following  are  some  of  the  vegetable  substances  recommended : 

(1)  Potato.  (5)  Turnip.  (9)  Onion.  (13)  Brazilnuts. 

(2)  Sweet  potato.  (6)  Radish.  (10)  Banana.  (14)  Apple. 

(3)  Carrot.  (7)  Salsify.  (n)  Coconut.  (15)  Pear  or  quince. 

(4)  Sugar-beet.  (8)  Parsnip.  (12)   Peanuts.  (16)  Pineapple. 

These  substances  may  be  extended  almost  indefinitely  and  are  very  useful  for 
making  preliminary  studies,  inasmuch  as  they  include  many  different  kinds  of 
chemical  substances.  The  writer  has  used  them  for  many  years.  They  should  be 
prepared  with  great  cleanliness,  especially  the  roots,  so  as  to  avoid  including  resistant 
spores.  Sterilization  is  an  easy  and  simple  process  if  these  substrata  are  free  from 
spores  when  the  steaming  begins.  Roots  and  tubers  should  be  selected  with  great 
care,  only  those  being  taken  which  are  sound  and  free  from  blemishes.  They  are  now 
to  be  washed  thoroughly  in  tap  water  with  scrubbing  and  then  rinsed  in  distilled 
water.  With  clean  hands  and  a  clean  knife  they  are  then  pared,  with  care  to  remove 
all  black  specks,  and  thrown  into  a  beaker  of  distilled,  filtered  or  boiled  water. 
Cylinders  of  the  proper  size  may  now  be  punched  with  a  clean  cork-borer  or  cut 
with  a  clean  sharp  knife  and,  after  the  xipper  part  has  been  slanted,  are  thrown 
into  another  beaker  of  distilled  water,  from  which  they  are  transferred  to  two  others 
before  they  are  finally  put  into  the  tubes.  It  is  not  necessary  to  soak  them  in  water 
over  night  or  in  antiseptic  solutions.  They  will  not  brown  by  oxidization  if  they  are 
kept  under  water  during  the  early  stages  of  preparation  and  are  steamed  as  soon  as 
they  are  placed  in  the  tubes,  i.  e.,  exposed  to  the  air.  They  may  be  put  into  the 
tubes  with  clean  fingers  or  by  means  of  a  pair  of  clean  forceps. 

On  these  different  media  observe  the  nature,  amount,  and  rapidity  of  growth 
(always  with  due  regard  to  the  air-temperature,  which  should  be  recorded) .  Carefully 
determine  whether  there  is  any  retardation  of  growth  at  first  and,  if  so,  to  what  it 
is  due,  so  that  more  exact  studies  may  be  made  subsequently  in  other  media.  Look 
for  gas-bubbles,  formation  of  acids  and  alkalies,  formation  of  hydrogen  sulphide, 
of  crystals,  of  stains,  of  odors,  destruction  of  starch,  disappearance  of  the  middle 
lamella,  softening  of  cellulose,  etc.  For  the  first  few  days  all  cultures  should  be 
examined  at  least  as  often  as  once  in  24  hours  and,  generally  speaking,  cultures 
should  not  be  discarded  until  after  the  sixth  or  eighth  week.  These  experiments 
should  be  repeated  a  number  of  times  and  the  student  should  avoid  drawing  a 
hasty  conclusion,  since  different  samples  of  potatoes,  carrots,  etc.,  vary  somewhat 
in  composition  and  will  at  times  give  slightly  varying  results  or  even  resxilts  which 
seem  to  be  contradictory,  e.g.,  a.  brown  pigment  in  some  instances  and  not  in  others. 


RAW    CULTURE    MEDIA. 


The  same  media,  and  as  many  other  sorts  as  are  available,  should  be  tested  raw 
in  sterile,  dry,  Petri  dishes  10  cm.  broad  and  2  to  3  cm.  deep.  For  this  purpose  the 
vegetables  are  prepared  as  follows  :  First,  select  sound,  clean  specimens,  especially 
avoiding  those  which  are  cracked  open  ;  next,  scrub  their  surface  thoroughly  under 
the  tap,  and  rinse  them  in  distilled  water.  They  are  now  soaked  5  or  10  minutes, 
or  even  20  minutes,  in  1:1000  water  solution  of  mercuric  chloride.  They  are  then 
removed  and  dried  with  or  without  a  preliminary  rinsing  in  sterile  water.  When 
dry  they  are  put  on  a  sterile  paper  or  plate,  are  cut  into  slices  about  1.5  to  2  cm. 
thick  with  a  cold  sterile  knife,  are  picked  up  with  sterile  forceps,  and  are  put  into 

the  Petri  dishes  in  pairs 
or  fours,  the  cover  being 
immediately  replaced. 
Enough  of  the  mercuric 
chloride  remains  on  the 
surface  to  inhibit  the 
growth  of  any  surface 
organisms  which  have 
not  been  killed  outright, 
and  experience  shows 
that  intruders  are  rarely 
dragged  over  the  cut 
surface.  The  slices  may 
be  inoculated  at  once 
or  after  36  hours  incuba- 
tion in  a  moist  chamber 
at  30°  C.,  or  48  hours  at 
25°  C.  The  latter  course 
is  preferable.  In  either 
case,  half  of  the  slices 
in  each  dish  must  be 
kept  uninoculated  for 


I 


Fig.  36.* 


comparison  (fig.  36).     This  method  is  well  adapted  to  the  study  of  various  soft-rot 
organisms  such  as  Bacillus  carotovorus,  B.  aroideiz,  B.  oleracetz,  etc. 

PI.ANT  JUICES  (WITH  AND  WITHOUT  THB  ADDITION  OF  WATER). 

(1)  Juice  of  the  host-plant.  (4)   Prune-juice. 

(2)  Potato-broth.  (5)  Orange-juice. 

(a)  With  sodium  hydrate.         (6)  Coconut-water  (unsteamed).t 
(fc)  Without  alkali.  (7)  Yellow  corn-meal  broth. 

(3)  Cabbage  or  cauliflower  broth.       (8)  Timothy-hay  infusion. 


*Fic.  36. — Iris-rhizome-rot  organism  grown  on  raw  carrot.  The  check  piece  is  unchanged,  the 
inoculated  piece  has  browned  and  softened.  Incubated  4  days  at  about  23°  C. 

fThis  is  removed  directly  from  the  nut  to  sterile  test-tubes  by  means  of  sterile  pipettes,  which 
are  useful  in  many  ways.  The  pipettes  should  be  dry-heated  and  kept  from  contamination  in  long, 
narrow,  covered  tin  boxes.  These  boxes  may  be  cylindrical  or  rectangular,  with  an  end  cover. 
The  upper  end  of  the  pipette  should  be  plugged  firmly  with  cotton  before  sterilization,  and  this 
should  be  pushed  in  a  short  distance  beyond  the  end,  so  that  when  the  finger  is  placed  on  the  end 
there  will  be  an  air-tight  union.  Scalpels,  etc.,  should  be  sterilized  in  shorter  boxes  of  the  same 
kind  (fig.  37). 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


These  fluids  are  only  a  few  of  many  that  may  be  used.  Some  of  them,  e.g., 
potato-broth,  require  special  care  in  preparation.  My  own  method  of  making  potato- 
broth  is  to  pass  the  clean  pared  potatoes  rapidly  through  a  grating  machine  and 

immediately  throw  the  pulp  into  the  re- 
quired quantity  of  distilled  water  (which  should 
be  twice  the  weight  of  the  potato).  The 
beaker  is  now  put  into  a  water-bath  and  the 
temperature  rapidly  raised  to  55°  C.  and  kept 
there  with  frequent  stirring  for  an  hour.  The 
pulp  is  now  filtered  from  the  fluid  and  the 
latter  is  immediately  put  into  the  steamer.  If 
the  steaming  is  long  delayed  the  broth  will  be 
dark  brown  (oxidizing  action  of  the  potato- 
enzyme  on  tannins  in  the  presence  of  air), 
and  if  the  temperature  rises  much  above 
60°  C.,  before  the  pulp  is  removed,  some  of 
the  starch  becomes  gelatinous  and  the  fluid 
will  not  filter. 

All  media  which  have  boiled  away  to  any 
Fig-  37.*  Fig.  38.  t         considerable  extent  must,  of  course,  be  made 

up  to  the  original  volume  or  weight  just  prior  to  final  sterilization. 

In  these  culture-fluids  observe  the  rapidity,  density,  and  persistency  of  the 
clouding ;  whether  the  clouding  is  simple  or  turbid  from  the  presence  of  zoogloese ; 
and  finally,  whether  it  is  uniform  in  all  parts  of  the  tube.  Note  the  character  of 
the  rim  and  pellicle,  if  any  are  formed,  and  how  soon  they  appear ;  also  the  amount, 
color,  and  general  appearance  of  the  precipitate.  The  amount  of  the  precipitate 
varies  greatly  with  different  media.  Its  quality  also  varies.  Sometimes  it  consists 
of  loose,  easily  separable  particles ;  in  other  cases  it  is  a  viscid  mass  which  rises  as 
a  rope-like  unit  when  the  tube  is  twirled  (fig.  38). 

Record  the  formation  of  acids,  alkalies,!  odors,  gas-bubbles,  stains,  crystals. 
Does  the  fluid  become  viscid  or  ropy  ?  Some  organisms  bring  about  this  condition 
quickly  in  a  variety  of  media,  e.  g.,  Bacterium  pericarditidis  (Bacillus  pyocyaneus 
pericarditidis),  others  rarely  or  never.  Precipitates  in  test-tube  cultures  vary  all  the 
way  from  a  scarcely  perceptible  trace  to  masses  a  centimeter  or  more  in  depth.  Do 
not  confound  chemical  precipitates  with  bacterial  growth.  Before  inoculation  always 
examine  media  in  test-tubes  for  presence  of  slight  precipitates  and  for  contaminat- 
ing organisms.  In  cultures  of  rapidly  growing  species,  at  optimum  temperatures, 
clouding  may  occur  in  less  than  twenty-four  hours ;  with  slow-growing  species,  and 

*Fio.  37. — Tin  box  for  holding  scalpels,  forceps,  etc.,  to  be  sterilized  by  dry  heat.  About  one- 
fourth  actual  size.  A  similar  tin  box  which  is  very  convenient  for  holding  sterile  pipettes  measures 
2  by  3  by  15  inches. 

tFic.  38.— Twirled  culture  of  the  olive-tubercle  organism  in  Uschinsky's  solution,  showing 
viscidity  of  the  precipitate  in  old  cultures. 

^Bacterial  ash  is  alkaline,  and  this  ash  must  be  carefully  washed  from  the  platinum  loop  in  dis- 
tilled water  each  time  before  it  is  used  to  transfer  drops  of  the  culture-fluid  to  litmus  paper.  The 
wire  must,  of  course,  be  re-flamed  after  washing. 


FILTERS. 


43 


o 


3 

Fig.  39.f 


when  the  medium  has  a  retarding  action,  it  may  not 
occur  until  after  two  or  three  weeks.  Of  course,  the 
rapidity  of  the  clouding  depends  to  a  considerable  extent 
on  the  size  of  the  loop  and  on  whether  the  inoculation 
was  from  a  young  or  old,  a  fluid  or  a  solid  culture. 

Among  other  tools,  the  student  should  be  provided 
with  five  platinum-iridium  wires  set  into  glass  handles, 
three  of  which  are  bent  at  the  free  end  into  loops  of  a 
definite  size,  i.  <?.,  with  an  inside  diameter  of  i,  2,  and 
3  mm.  These  are  made  by  bending  around  wires  of  the 
given  size,  and  will  enable  one  to  measure  out  approxi- 
mately uniform  quantities  of  fluids  and  solids.  Smaller 
quantities  may  be  transferred  on  the  extreme  tip  of  a 
straight  platinum  needle.  It  is  also  convenient  to  have 
a  platinum*  needle  bent  at  the  end  into  a  short  hook 
(see  fig.  39).  In  comparing  rates  of  growth  in  fluid 
cultures  it  is  best  to  inoculate  them  from  other  fluid 
cultures  of  a  given  age  and  not  from  solids. 

If  there  is  any  reason  to  think  that  boiling  changes 
the  nature  of  any  of  these  fluids,  they  should  be  steril- 
ized cold  by  forcing  them  through  a  Chamberland  or 
Berkefeld  filter.  The  Chamberland  has  the  finer  pores, 
the  Berkefeld  filters  quicker.  The  simplest  way  of  using 
such  a  filter  is  that  first  described  by  Dr.  Theobald 
Smith,  viz,  to  put  the  fluid  inside  and  force  it  out  by 
means  of  clean  compressed  air.  For  this  purpose  select 
a  flat-bottomed  cylindrical  glass  vessel  (a  round-bottomed 
one  is  less  convenient,  but  may  be  set  into  a  hole  bored 
in  a  block  of  wood)  of  a  larger  diameter  and  5  or  10 
centimeters  longer  than  the  bougie,  which  should  be 
clean  (previously  unused),  but  washed  by  having  had 
some  liters  of  distilled  or  filtered  water  forced  through 
it.  Wrap  the  nipple-end  of  the  filtering  cylinder  firmly 
with  clean  cotton  for  a  distance  of  5  or  10  cm.  down. 
Thrust  the  wrapped  bougie  into  the  glass  vessel  securely, 
so  that  only  the  nipple  and  the  cap  or  shoulder  projects. 
The  top  of  the  bougie  should  also  be  wired  so  that  it 
can  not  possibly  slip  down  during  the  filtering.  This 
apparatus  should  now  be  sterilized  by  putting  it  into 
the  dry  oven  for  two  hours  at  145°  C.  Wrap  in  clean 
Manila  paper  and  heat  at  the  same  time  a  large  cotton 
plug,  i.  e.,  one  which  has  been  made  to  fit  the  mouth 


*Platinum^iridium  is  preferred  to  pure  platinum  because  it  bends  less  easily.  The  wire  used  by 
the  writer  has  a  diameter  of  0.48  nun.  The  alloy  as  usually  found  on  the  market  is  said  to  contain 
about  10  per  cent  of  indium,  sometimes  less,  but  never  more.  The  wire  shown  in  fig.  39  was  made 
to  order  and  contains  20  per  cent  iridium. 

tFic.  39. — Platinum-iridium  wires  set  into  glass  rods,  for  bacteriological  work.  I,  needle; 
2,  hook ;  3,  one-millimeter  loop ;  4,  two-millimeter  loop ;  5,  three-millimeter  loop.  The  size  of  this 
wire  is  about  one  fifty-fifth  inch. 


44 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


of  the  cylindrical  glass  vessel.  Wheii  sterilized  and  ready  for  use,  select  a  piece 
of  rubber  cloth  10  or  15  cm.  in  diameter,  cut  a  small  slit  in  its  center  and  draw 
it  over  the  nipple  of  the  bougie  to  protect  the  cotton  from  accidental  wetting  and 
the  filtered  fluid  from  consequent  possible  contamination.  Now  pour  the  fluid 
into  the  bougie  (if  one  with  a  large  neck  has  been  selected  this  will  not  be  difficult, 
especially  if  a  small  funnel  is  used  and  this  is  kept  from  close 
contact  on  one  side  by  means  of  a  small  wire,  sliver,  or  bit  of 
paper),  and  connect  the  nipple  with  the  outflow-tube  of  the 
compressed-air  pipe  by  means  of  an  extra-thick  rubber  tube, 
which  should  be  securely  wired  at  each  end,  and  turn  on  the 
compressed  air  cautiously.  Fluids  which  are  not  colloidal 
usually  filter  very  readily  with  a  pressure  of  15  or  20  pounds 
per  square  inch. 

The  filtering  should  always  be  done  slowly  with  a  mini- 
mum pressure  in  order  to  avoid  the  possibility  of  forcing  small 
organisms  through  the  walls  of  the  filter.  With  heavy  pressure 
this  sometimes  occurs  when  no  cracks  are  detectable  in  the 
bougie.  When  the  desired  quantity  of  fluid  has  been  filtered 
(fig.  40)  cut  off  the  air-blast,  disconnect  the  tube,  tilt  the  cylinder 
as  much  as  possible,  remove  the  bougie,  and  substitute  the 
sterile  cotton  plug.  The  fluid  should  now  be  transferred 
immediately,  in  5  or  10  cc.  portions,  to  sterile  cotton-plugged 
test-tubes  by  means  of  sterile  pipettes.  The  removal  of  the 
bougie  and  the  transfer  of  the  fluid  should  be  done  in  clean 
still  air,  under  a  hood  or  in  a  special  culture-room.  The  tubes 
should  not  be  used  for  several  days,  i.  e.,  time  should  be  given 
for  contaminations  to  show  themselves,  but  if  proper  care  has 
been  exercised  there  should  be  very  few  contaminations  or 
none  at  all.  A  pressure  much  greater  than  20  pounds  per 
square  inch  may  be  obtained  by  means  of  steam-pumps  or  by 
use  of  cylinders  of  compressed  air,  oxygen,  or  carbon  dioxide, 
and  this  is  sometimes  necessary  for  colloidal  substances,  but 
should  be  used  cautiously.  These  cylinders  may  be  had  from 
the  Eagle  Oxygen  Company,  New  York.  One  of  the  most 
convenient  filters  on  the  market  is  that  shown  in  fig.  41.  It 
was  designed  by  Roux  and  is  made  by  Maison  Wiesnegg 
(P.  Lequeux),  Paris.  It  is  well  made,  very  durable,  quickly 
sterilized,  and  easily  operated  if  one  can  command  an  air-blast 
or  other  gas-pressure  of  2  or  3  atmospheres. 

Chamberland  bougies  ought  not  to  be  used  continuously  for  more  than  three 
days.     They  should  then  be  removed  and  baked  for  two  hours  at  145°  C.  (or  at  the 

*FiG.  40. — Simple  method  of  obtaining  small  quantities  of  sterile  fluids  by  means  of  the 
Ohamiberland  filter.  The  other  end  of  the  rubber  tube  is  wired  securely  to  the  outflow  pipe  of  the 
compressed-air  system  and  the  fluid  is  forced  from  the  inside  of  the  filter  out.  This  method  was 
first  described  and  figured  by  Dr.  Theobald  Smith.  About  one-fourth  actual  size. 


Fig.  40* 


FILTERS. 


45 


temperature  of  an  oven  in  which  bread  is  baked).    The  reason  for  this  lies  in  the  fact 
that  in  three  or  four  days  time  certain  small  organisms  are  able  to  grow  through  the 

walls  of  the  filter  and  make  their  appear- 
ance in  the  filtered  fluids  on  the  other 
side.  Persons  who  never  bake  their 
water-filters  rest  in  unwarranted  secu- 
rity. The  bougies  must  also  be  handled 
with  great  care  and  inspected  carefully 
after  each  baking  for  the  appearance  of 
minute  cracks.  To  detect  cracks,  im- 
merse the  tube  in  water  and  blow  into 
it.  Clogged  filters  should  be  sent  to 
the  firers  of  china,  where  they  may  be 
purified  by  heating  to  dull  redness. 

ANIMAL  FLUIDS. 
BEEF-BROTH. 

(a)  Acid,  neutral,  and  alkaline. 

(b)  The  same,  with  addition  of  0.5 
per  cent  c.  p.  sodium  chloride  and  i  per 
cent  peptone  (Witte's  peptonum  siccum, 
Merck's    brown    peptone,    Savory    & 
Moore's  brown  peptone,  etc.).     This  is 
ordinary  peptonized  beef-broth. 

Examine  as  in  case  of  plant  juices. 
The  term  peptone,  as  it  occurs  in  bac- 
teriological literature,  usually  means 
commercial  peptone,  which  is  a  mix- 
ture of  true  peptone  and  various  pro- 
teoses  or  albumoses.  It  is  therefore 
generally  best  to  specify  just  what  pep- 
tone is  used.  The  writer  now  gen- 
erally uses  Witte's  dry  white  peptone. 
Savory  &  Moore's  brown  peptone  from 
flesh  is  very  good  for  some  purposes. 

*Pic.  41. — Dr.  Roux's  pressure-filter,  made  by  Maison  Wiesnegg  (P.  Lequeux),  Paris.  The 
working  capacity  of  this  filter  is  about  1.3  liters.  The  principal  parts  are:  A,  tube  for  connection 
with  compressed-air  system;  B,  cut-off;  C,  cover  held  in  place  (by  strong  bolts;  D,  central  reser- 
voir; E,  cut-off;  F,  screw  collar  which  holds  the  bougie  in  place;  G,  heavy  •metal  cylinder  surround- 
ing the  bougie;  H,  cut-off,  which  is  closed  of  course  when  the  apparatus  is  in  use;  I,  funnel 
through  which  G  and  D  are  filled ;  K,  device  for  sterilizing  the  interior  of  the  apparatus  by  steam 
under  light  pressure  (it  consists  of  a  copper  chamber  partly  full  of  distilled  water,  to  the  bottom 
of  which  the  Bunsen  flame  is  applied;  the  chamber  may  be  unscrewed  and  removed);  L,  button 
which  is  unscrewed  to  fill  the  chamber  with  water  (in  its  center  is  a  steam  safety  valve  acting 
under  feeble  pressure)  ;  M,  valve  which  cuts  the  steam-generator  out  of  the  general  circulation 
when  fluids  are  being  filtered;  N,  tripod-top  on  which  the  apparatus  turns  freely.  Height,  33  inches. 


46 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


Milk. — Milk  from  a  clean  dairy  and  free  or  nearly  free  from  cream  should  be  selected 
for  use.  If  some  cream  remains  it  may  be  filtered  out  or  removed  by  the  centrifuge 
(fig.  43).  The  milk  should  not  be  acid  to  the  taste  and  should  not  contain  formal- 
dehyd  or  other  antiseptic  substances  which  milk-dealers  sometimes  add  to  dirty 
milk  to  improve  its  keeping  qualities.  It  should  be  steamed  in  wire-crates  1 5  min- 
utes at  100°  C.  on  each  of  four  consecutive  days  (10  cc.  portions  in  test-tubes),  and 
should  not  be  used  until  at  least  a  week  after  the  last  steaming.  Such  milk  should 
titrate  -+-  12  to  +  17  or  thereabouts  with  sodium  hydrate  and  phenolphthalein.  Milk- 
cultures  should  be  kept  under  observation  at  least  six  or  eight  weeks. 

Observe  in  particular : 

(a)  Separation  of  the  casein  without  the  develop- 
ment of  any  acid,  indicating  the  presence  of 
the  lab,  or  rennet,  ferment.  The  milk  usually 
becomes  more  alkaline. 

(6)  Saponification  of  the  fat.  The  fluid  becomes 
transparent  without  any  precipitation  of  casein; 
but  the  caseinogen  may  be  thrown  down  sub- 
sequently by  acidifying  the  clear  liquid. 

(c)  Ropiness.    The  fluid  becomes  viscid,  and  strings 

when  touched.  This  viscidity  is  sometimes  so 
great  that  an  entire  pail  of  milk  may  be  in- 
verted without  immediate  loss  of  its  contents. 
See  striking  figures  in  Ward's  papers  ( '99  and 
'oi,  Bibliog.,  XLVII). 

(d)  Formation  of  acids.    This  occurs  with  or  with- 

out evolution  of  gas,  and  usually  with  the  final 
separation  of  the  whey  from  the  casein  at  room 
temperatures  or  on  boiling.  Boil  if  necessary. 

(e)  Re-solution  of  precipitated  casein  ( trypsin  fer- 

ment); formation  of  crystals  (tyrosin,  leucin, 

etc. ). 

(/)  Gelatinization  of  old  cultures.     Milk  alkaline. 
(g )  Changes  in  smell,  color,  and  taste. 

In   using   milk  it   should  not  be  for- 
gotten that  anaerobes  are  sometimes  pres- 
ent (Theobald  Smith)  and  also  organisms 
of  the  dunghill  which  will  grow  only  at 
temperatures  above  40°  C.    Very  resistant 
spores  of  aerobic  species,  growing  at  tern- 
Fig.  42*  peratures  below  40°  C.,  are  present  also 
sometimes,  especially  in  dirty  milk,  and  the  milk  is  then  difficult  to  sterilize. 

Several  experiments  made  by  the  writer  with  milk  from  Washington  dairies 
have  shown  that  Franz  Lafar's  statement  in  Technische  Mykologie,  Bd.  I,  p.  189, 
while  probably  true  for  the  milks  which  he  tested,  is  not  true  when  stated  as  a 
general  proposition.  In  brief,  this  statement  is  that  nine  out  of  ten  milks  are  not 


*Fic.  42.— Section  of  Arnold  steam  sterilizer.  Water  enters  ,the  .double  'bottom  through  a  few 
small  openings  indicated  by  two  arrows  in  the  water-pan.  The  other  arrows  show  movement  of 
the  steam.  In  this  form  the  outer  jacket  (of  copper)  is  lifted  off  to  put  in  or  remove  media. 


STERILIZATION. 


47 


sterilized  by  steaming  twenty  to  thirty  minutes  on  three  consecutive  days,  but  will 
develop  bacterial  growths  when  put  into  the  thermostat.     If  such  were  really  the 
case,  milk  would  be  one  of  the  worst  of  culture-media  instead  of  one  of  the  best 
The  general  experience  of  bacteriologists   is  not  in  accord  with  this  statement. 
Occasionally,  in  my  own  experience,  a  single  steaming  of  five  or  ten  minutes  has 

r  sufficed    to    sterilize    milk 

completely,  at  least  so  far  as 
relates  to  organisms  which 
grow  aerobically  and  at  tem- 
peratures under 4o°C.  Such 
milks  have  remained  un- 
changed for  two  or  three 
months  at  room  tempera- 
tures (20°  to  25°  C.),  and 
also  in  the  thermostat  at 
blood  heat.  For  anaerobes, 
or  organisms  which  will 
grow  only  at  temperatures 
above  40°  C.,  I  have  not 
tested. 

One  possible  source  ol 
error  in  the  use  of  steam 
for  sterilization  is  ignorance 
of  the  exact  temperature  of 
the  steam-chamber.  Every 
steam-sterilizer  should  have 
a  hole  punched  through  the 
top,  into  which  is  fitted  a 
cork  through  which  a  ther- 
mometer projects  into  the 
chamber.  In  this  way  may 
be  determined  beyond  doubt 
for  just  how  many  minutes 
the  media  has  been  exposed 
to  steam  at  100°  C.  The 
Arnold  steam  -  sterilizer, 
which  is  one  of  the  best,f 
is  greatly  improved  by  this 
simple  device  (fig.  42  and 
pi.  6).  In  this  sterilizer  there  is  a  double  bottom  under  the  water-pan.  The  lower 
bottom  is  in  contact  with  the  Bunsen  flame.  Through  small  holes  in  the  upper 

*Fic.  43. — Improved  Lautenschlager  centrifuge.  Capacity,  540  cc.  Revolutions  per  minute, 
3,000  to  4,000.  It  requires  about  3  horsepower  to  run  the  apparatus  at  this  high  speed.  About  one- 
eleventh  natural  size. 

fThis  remark  does  not  apply  to  the  Arnold  combination  steamer  and  dry  oven,  which  can  not 
be  recommended. 


Fig  43.« 


48 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


bottom  the  water  drips  to  the  lower  bottom  and  is  quickly  converted  into  steam 
which  streams  through  a  central  chimney  into  the  bottom  of  the  sterilizing  chamber. 
The  latter  has  two  walls,  with  a  considerable  air-space  between,  open  at  the  bottom. 
The  streaming  steam  passes  over  the  top  of  the  inner  wall  downward  into  this  air- 
space and  escapes  into  the  pan  as  condensation  water.     Theoretically  this  is  a  very 

perfect  sterilizer,  and  it  is  so  in  prac- 
tice when  new,  but  not  infrequently 
it  leaks,  and  sometimes  the  openings 
in  the  upper  bottom  are  too  large  or 
become  clogged  by  mud.  When  in 
perfect  working  order  it  takes  only  a 
few  minutes  to  get  a  temperature  of 
100°  C. 

Tubes  should  always  be  steamed 
in  wire-crates  (fig.  44)  so  that  the 
streaming  steam  may  have  full  access 
to  all  parts.  Tubes  of  media  steamed 
in  cans  or  beakers  often  spoil.  They 
seem  to  retain  a  cushion  of  air  about 
them  which  interferes  with  the  action 
of  the  steam. 

Litmus  milk. — Litmus  milk  of  a 
good  quality  may  be  made  by  dissolv- 


Fig.  44  * 


ing  Merck's  dry,  lime-free  c.  p.  blue 
litmus  to  saturation  in  distilled  water 
(1:15)  and  then  adding  one  part  of  this  blue  fluid  to  each  fifty  parts  of  milk.  The 
milk  should  be  a  deep  lavender  color.  Much  inferior  litmus  is  on  the  market. 
Large  use  should  be  made  of  this  fluid.  In  addition  to  observations  under  "Milk," 
note  how  rapidly  the  litmus  reddens,  blues,  or  becomes  reduced,  and  how  soon  the 
color  returns.  Will  it  return  at  once  on  steaming  the  culture? 

Rice  cooked  in  milk. — (One  or  two  grains  to  10  cc.  in  each  test-tube).  This  is 
useful  for  study  of  some  chromogens. 

Lpejfler^s  solidified  blood-serum. — Observations  under  this  and  the  following 
heads  are  the  same  as  for  gelatin  slant  cultures.  The  plant  bacteriologist  must  in 
general  obtain  blood-serum  from  the  animal  bacteriologist.  The  solidified  serum 
may  also  be  used  plain,  *.  e.,  without  the  addition  of  grape-sugar. 

Egg-albumen.— This  is  solidified  and  used  in  the  same  way  as  blood-serum. 
The  end  of  the  egg  from  which  the  albumen  is  poured  must  be  thoroughly  flamed 
before  it  is  broken,  and  care  must  be  used  in  the  transfer  to  test-tubes  so  as  to 
exclude  air-borne  germs  as  far  as  possible,  otherwise  the  sterilization  will  be  difficult. 
The  albumen  of  eggs  may  be  cut  with  sterile  scissors. 


*Fic.  44.— Wire-crate  for  holding  tubed  culture-media  which  is  to  be  steamed.     About  two- 
fifths  actual  size.     A  tuft  of  cotton  on  the  bottom  prevents  the  breaking  of  tubes. 


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MEDIA.  49 

Egg-yolk. — This  is  poured  into  test-tubes  and  solidified  in  a  slanting  position 
by  heat  (80°  C.),  or  the  egg  may  be  boiled  hard  and  the  yolk  cut  with  a  sharp 
knife  and  transferred  to  sterile  Petri  dishes.  If  desired,  the  yolk  and  white  may 
be  mixed  before  solidifying,  i.  e.,  by  shaking  the  egg  vigorously  before  breaking 
the  shell. 

SYNTHETIC  MEDIA  AND  OTHER  SPECIAL  MEDIA. 

The  student  should  try  the  following  media.  He  should  also  invent  media  to 
suit  special  cases.  The  kinds  of  media  I  have  in  mind  are  the  opposite  of  universal, 


Fig.  45  * 

to- wit,  such  as  will  favor  the  growth  of  some  organisms  while  preventing  that  of 
others.  The  acid  phosphates  and  many  other  substances  are  useful  for  this  purpose. 
The  field  is  comparatively  new,  and  much  is  to  be  learned  by  careful  experimenting. 

1.  Dunham's  solution. 

2.  Peptone-water   (i  or  2  per  cent)    with   addition  of  various    carbohydrates, 
acids,  etc. 

3.  Sugar-free  beef  bouillon  with  Witte's  peptonum  siccum  (for  the  indol  test). 

4.  Cohn's  solution. 

5.  Uschinsky's  solution. 


*Kig.  45. — Oven  for  solidifying  and  sterilizing  blood-serum,  nutrient  starch-jelly,  silicate-jelly, 
etc.,  at  temperatures  below  100°  C.  When  in  use  the  temperature  is  controlled  by  means  of  a 
Tollen's  thermo-regulator  (see  fig.  35). 


50  BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 

6.  Uschinsky's   solution  with  various  carbon  compounds  substituted  for  the 
glycerin  (fermentation  tubes). 

7.  Fraenkel  and  Voge's  solution. 

8.  Raulin's  solution. 

9.  Fermi's  solution. 

10.  Water  (distilled),  1,000, ooomg.;  dipotassium  phosphate,  2,000  mg.;  ammonium 
phosphate,  100  mg.;  magnesium  sulphate,  100  mg.;  sodium  acetate,  5,000  nig. 

1 1 .  Same,  with  the  carbon  compound  changed,  e.  g. ,  with  sodium  formate  substi- 
tuted for  sodium  acetate.     Sodium  formate  and  phenol phthalein   may  be  added  also 
to  bouillon  or  agar  (2  per  cent)  for  observations  during  the  early  stages  of  growth, 
some  organisms  reddening  this  medium  promptly  by  decomposition  of  the  sodium  salt. 
(See  a  recent  paper  by  Omelianski). 

12.  Nutrient  starch-jelly  for  study  of  diastasic  action.    (SeeProc.Am.  Asso.  Adv. 
Sci.,  1898,  p.  411,  or  Centralb.  f.  Bakt.,  2te  Abt.,  Bd.  V.,  p.  102.) 

One  gram  of  starch  is  rubbed  up  -with  a  sterile  glass  rod  in  10  cc.  of  the  sterile  nutrient  fluid 
(Uschinsky's  solution,  etc.),  placed  in  a  slanting  position,  in  test-tubes,  and  solidified  in  a  blood- 
serum  oven  (fig.  45)  or  in  the  .top  of  a  steamer  with  the  vents  left  open.  There  should  be  several 
heatings  of  two  hours  each  to  insure  sterilization.  The  .temperature  should  not  exceed  93°  C.  nor 
fall  much  below  85°  C.  Sterilization  is  rendered  much  easier  if  the  starch  is  prepared  in  a  cleanly 
way.  The  only  difficulty  the  writer  has  experienced  is  in  the  formation  of  a  thin  film  of  semi- 
opaque  solidified  starch  on  the  walls  of  the  tubes  above  the  slant.  This  often  cracks  off,  however, 
during  the  heatings,  and  is  largely  obviated  by  placing  the  tubes  in  a  slanting  position  before  the 
starch  is  rubbed  up  in  the  fluid,  taking  care  to  soil  the  walls  above  the  slant  surface  as  little  as 
possible  during  the  operation.  The  potato-starch  is  prepared  as  follows : 

One-half  bushel  of  large  smooth  >potatoes  are  scrubbed,  and  the  black  specks  dug  out;  they  a-re 
then  soaked  for  45  minutes  in  I  :iooo  mercuric-chloride  water.  Meanwhile  the  hands  are  scrubbed 
clean  and  given  a  five  minutes  washing  in  the  mercuric-chloride  water.  The  tubers  are  now  rinsed 
in  sterile  water,  pared  deeply,  grated  as  for  potato-broth,  and  thrown  into  beakers  containing  sev- 
eral liters  of  distilled  water,  where  the  pulp  is  worked  over  with  the  hands  to  liberate  as  much 
starch  as  possible.  The  starchy  water  is  now  removed  from  tine  pulp  by  passing  it  through  several 
folds  of  surgeon's  gauze,  squeezing  out  of  the  pulp  as  much  of  the  fluid  as  possible.  When  the 
starch  has  settled  the  brownish  fluid  and  floating  fragments  are  poured  off  or  decanted,  and 
fresh  distilled  water  is  added.  The  smaller  fragments  of  cell-wall,  etc.,  are  then  removed  by  forc- 
ing the  starch  (stirred  up  in  water)  through  a  moderately  fine-meshed  towel  (not  too  fine)  with 
gentle  hand-rubbing,  into  another  beaker.  Most  of  the  medium-sized  and  finer  starch-grains  pass 
through,  leaving  in  the  towel  the  coarser  grains  and  those  fragments  of  cell-wall  which  passed 
through  the  coarser  meshes  of  the  surgeon's  gauze.  The  purified  starch  is  now  allowed  to  stand 
for  about  a  week  in  the  >ice-t>ox  in  distilled  water  (3  liters  or  more  per  'beaker  or  jar).  The  water 
is  siphoned  off  twice  a  day  at  first,  and  afterwards  once  a  day,  the  starch  being  stirred  up  thor- 
oughly every  time  fresh  water  is  added.  Finally  the  starch  is  drained  very  free  from  water,  scooped 
out  with  sterile  spoons  or  spatulas,  placed  in  uncovered  sterile  Petri  dishes,  and  dried  in  the 
blood-serum  oven  at  56°  C.,  the  cover  being  raised  an  inch  (on  corks)  to  let  the  moisture  out. 
One-half  bushel  of  sound  potatoes  should  yield  from  400  to  500  grams  of  air-dry  aseptic  starch. 

Potato  starch  has  been  selected  because  it  is  easy  to  prepare,  but  other  starches  might  yield  in- 
teresting results.  Bacteriologists  now  pay  great  attention  to  the  fermentation  of  sugars,  but  thus  far 
very  little  consideration  has  been  given  to  the  action  of  bacteria  on  starches  and  celluloses.  What- 
ever starches  are  used,  they  should  be  prepared  in  the  laboratory,  under  aseptic  conditions,  so  as  to 
exclude  spore-bearing  organisms. 

13.  Starch-jelly  with  addition  of  various  sugars,  gums,  and  alcohols  (for  study 
of  organisms  having  little  or  no  action  on  starch). 

14.  Tubes  of  slant  nutrient  agar  (  +  15  of  Fuller's  scale)  with  varying  amounts 
of  c.  p.  glycerin,  2  to  10  per  cent  or  more. 


SYNTHETIC    MEDIA.  51 

15.  Tubes  of  10  cc.  slant  agar  with  10,  20,  and  30  grams  of  grape-sugar. 

1 6.  The  same,  with  the  same  amounts  of  cane-sugar. 

17.  Gelatin  with  cane-sugar,  varying  amounts. 

18.  Gelatin  with  malic  acid.     (17  and  18  may  be  combined.) 

19.  Gelatin  plates  with  soluble  starch  and  i  per  cent  potassium  iodide  and  with 
or  without  i  per  cent  potassium  nitrate.     Try  a  mixture  of  the  pear -blight  organism 
and  B.  coli.     Can  the  colonies  be  distinguished  in  this  way  using  the  nitrate? 

20.  Agar  plates  with  various  sugars  and  the  addition  of  calcium  carbonate,  or 
zinc  carbonate,  for  detection  of  acid-forming  colonies.    ('91,  Beyerinck,  Bibliog.,  XX.) 

21.  Silicate-jelly.     See  p.  36.     Known  also  as  silica-jelly, 

22.  Nitrate  bouillon  (+  15  bouillon  with  i  per  cent  potassium  nitrate). 

23.  Triple-distilled  water  and  nutrient  mineral  substances  free  from  nitrogen. 
The  same,  with  addition  of  potassium  nitrate.     The  same,  with  other  nitrogen  foods, 
e.g.,  sodium  asparaginate. 

24.  Bouillon  with  lead  acetate. 

25.  Bouillon  with  neutral  red. 

26.  Salt  bouillon,  i.  e.,  +   15  bouillon  with  varying  amounts  of  c.  p.  sodium 
chloride  (i  to  5  percent). 

27.  Standard  peptonized  bouillon  with  varying  amounts  of  sodium  hydrate  (from 
-f-  40  to  —  40)  for  determining  the  optimum  reaction  and  the  tolerated  range  of  acidity 
and  alkalinity. 

Synthetic  media  may  be  varied  indefinitely  to  fit  special  cases  and  are  often 
extremely  useful  as  differential  tests.  They  have  frequently  been  condemned  because 
some  particular  organism  has  not  grown  well  in  them.  The  very  fact  of  feeble 
growth  or  of  no  growth  is,  however,  a  matter  of  interest,  and  not  infrequently  a 
means  of  distinguishing  organisms  which  resemble  each  other  in  many  particulars. 
The  value  of  such  media  becomes  apparent  at  once  when  a  number  of  organisms 
are  compared.  Synthetic  media  afford  more  exact  methods  of  research  than  do  the 
common  media,  and  their  value  must  increase  rather  than  diminish  as  time  goes 
on.  (Consult  Grimbert  in  Archives  de  Parasitologie,  T.  I,  pp.  191-216.)  It  does  not 
follow,  however,  that  the  common  media  should  be  at  once  abandoned.  Festina 
lente  is  a  good  rule.  The  formulae  for  some  synthetic  media  are  given  under 
"  Formulae."  For  others  see  various  text-books  and  the  papers  cited  in  the  Bibli- 
ography under  XVI,  XVII,  XVIII,  XXV,  etc. 

RELATION  TO  FREE  OXYGEN. 

(/)  Surface  and  deep  grmvths. — Note  the  behavior  of  deep  stabs  in  tubes  of 
recently  steamed  gelatin  and  agar,  or  of  the  colonies  in  shake-cultures  of  gelatin 
and  agar  which  are  protected  from  the  free  action  of  air  by  pouring  into  the  tubes 
as  soon  as  solidified  another  tube  of  gelatin  or  agar  in  the  surface  layers  of  which, 
as  an  additional  precaution,  some  active  aerobe  may  be  grown,  e.  g.,  Bacillus  sub- 
tilts.  Observe  also  the  relative  rate  of  growth  of  buried  and  surface  colonies  in  plate 
cultures,  growth  under  sterile  mica  plates,  etc.  Of  course,  whether  an  organism 
will  or  will  not  grow  under  the  conditions  mentioned  depends  often  to  a  large  extent 
on  the  composition  of  the  culture  medium.  It  might  be  able  to  respire  in  the  pres- 
ence of  grape-sugar  or  cane-sugar,  but  not  when  milk-sugar  or  glycerole  is  substi- 


52  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

tuted.  It  will  not  do  to  conclude  that  an  organism  is  a  strict  aerobe  until  it  has 
been  tested  anaerobically  in  the  presence  of  a  variety  of  carbon  foods  with  uni- 
formly negative  results.  One  who  has  had  some  experience  may  often  give  a  shrewd 
guess  as  to  behavior  in  fermentation-tubes  by  carefully  noting  the  growth  of  buried 
and  surface  colonies  in  ordinary  media. 

(2)  Fermentation-tubes. — The  fluids  may  be  Uschinsky's  solution  (without  the 
glycerin  unless  this  is  the  carbon  compound  to  be  tested);  peptone  water  (2  per 
cent  Witte's  peptone  with  0.5  per  cent  sodium  chloride);  and  filtered  tap  water,  or 
sugar-free  beef  bouillon  with  addition  of  i  per  cent  Witte's  peptone  (preferably  for 
most  purposes  this  latter  fluid).  The  substances  to  be  tested  (which  should  be 
chemically  pure  or  as  nearly  so  as  possible)  are  grape-sugar,  fruit-sugar,  cane-sugar, 
milk-sugar,  galactose,  maltose,  dextrin,*  mannit,  dulcit,  raffinose,  glycerin,  ethyl 
alcohol,!  methyl  alcohol,  acetone,  ammonium  lactate,  ammonium  tartrate,  asparagin, 
sodium  asparaginate,  urea,  etc.  One  to  5  per  cent  of  the  various  sugars,  etc.,  may 
be  used  ;  2  per  cent  is  a  good  quantity. 


Fig.  464 

Observe  carefully  what  substances  induce  clouding  in  the  closed  end  and 
whether  any  gas  is  produced.  Test  from  time  to  time  for  acids.  The  relative  vigor 
of  growth  in  the  open  end  should  also  be  noted.  Does  growth  stop  in  the  U  with 
a  sharp  line  of  demarcation?  Does  the  addition  of  calcium  carbonate  reduce  or 
prevent  the  formation  of  gas  or  favor  growth  in  any  way?  Is  the  reaction  in  the 
closed  end,  as  the  result  of  growth,  different  from  that  in  the  open  end  ?  Pipette 
out  all  the  fluid  from  the  open  end,  determine  its  reaction  to  litmus,  and  then  test 
the  reaction  of  that  which  remains.  How  is  the  difference,  if  any,  accounted  for  ? 
If  growth  finally  ensues  in  the  closed  end,  is  there  any  reason  for  thinking  it  due 
to  absorbed  air?  How  can  this  be  determined  ? 

It  should  be  remembered  that  often,  after  a  time,  air  is  absorbed  into  the  closed 
end  of  fermentation-tubes  and  may  lead  to  confusing  results.  For  this  reason, 
if  they  have  stood  on  the  shelf  any  length  of  time  after  sterilization,  they  should 
be  re-steamed  and  the  bubble  of  air  tilted  out  before  they  are  inoculated.  They 


*The  dextrin  should  be  freely  soluble  in  cold  water  and  should  not  give  any  red  reaction  with 
iodine — t.  e.,  should  be  free  from  amylo-dextrin  (erythro-dextrin).  Such  dextrin  is  hard  to  procure. 

tThis  and  the  next  four  should  be  added,  after  sterilization,  by  means  of  sterile  pipettes.  The 
ammonium  salts  may  be  obtained  in  a  sterile  condition  without  loss  of  ammonia  by  dissolving 
10  grams  in  200  cc.  of  water  and  forcing  this  through  a  Chamberland  filter  into  a  sterile  flask, 
from  which  the  proper  quantity  may  be  pipetted  into  the  culture  medium  after  sterilization. 

\  FIG.  46. — 'Wooden  carrier  for  fermentation-tubes,  the  flanging  base  being  held  under  the 
grooves.  Muoh  reduced. 


RELATION    TO   OXYGEN. 


53 


should  be  disturbed  as  little  as  possible  after  inoculation,  and  especially  all  tiltings 
or  rough  jarring  should  be  avoided.  They  may  be  carried  in  a  wooden  rack  (fig.  46). 
All  culture-media,  whether  inoculated  or  not,  should  be  protected  from  light. 

Figs.  47,  48,  49  show  fermentation-tubes  in  actual  use. 

The  pattern  of  fermentation-tube  preferred  by  the  writer  is  that  slight  modi- 
fication of  Einhorn's  tube  designed  by  Dr.  Theobald  Smith  (see  Wilder  Quarter 
Century  Book).  The  tubes  may  be  had  from  Emil  Greiner,  New  York.  Certain 


Fig.  47* 


Fig.  48.t 


Fig.  494 


forms  of  tubes  should  not  be  used.  One  of  these,  a  short,  thick  tube  with  a  wide 
U,  in  use  in  some  laboratories  in  this  country,  allows  air  to  pass  readily  into  the 
closed  end  and  is  entirely  worthless.  A  sample  tube  of  this  sort  was  filled  with 

*Fic.  47. — Fermentation-tube  with  Bacillus  tracheifhilus,  showing  absence  of  gas  and  uniform 
clouding  in  open  and  closed  end  in  the  presence  of  grape-sugar.  The  fluid  consisted  of  water,  400; 
Savory  &  Moore's  peptone,  4;  sodium  chloride,  i;  c.  p.  grape-sugar,  2;  saturated  solution  carbonate 
of  soda  (20°  C),  20  drops,  »'.  e.,  enough  to  render  the  fluid  slightly  alkaline  to  litmus. 

tFiG  48. — Fermentation-tube  with  Bacillus  tracheiphilus,  showing  inability  of  organism  to  grow 
anaerobically  with  glycerin  as  the  carbon  food.  Fluid,  distilled  water  with  I  per  cent  Witte's  pep- 
tonum  iiccum  and  I  per  cent  Schering's  c.  p.  glycerin.  Copious  growth  in  open  end  and  in  outer 
part  of  U ;  none  in  the  closed  end. 

jFic.  49. — Fermentation-tube  of  cane-sugar  peptone  water  inoculated  with  a  white,  gas-forming 
organism  plated  from  a  spot  disease  of  sisal  hemp.  The  total  amount  of  gas  produced  and  its  rate 
of  evolution  at  20°  to  23°  C.  are  indicated  by  marks  on  the  closed  end  of  the  tube. 


54 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


beef-bouillon  and  steamed  every  twenty-four  hours  for  seven  or  eight  days,  a  large 
bubble  being  tilted  out  each  time  and  appearing  just  as  regularly  during  the  next 
steaming.  Naturally,  no  strict  anaerobe  would  grow  in  such  a  tube  and  every 
aerobe  would  appear  to  be  a  facultative  anaerobe.  The  neck  of  the  fermentation- 
tube  should  be  as  narrow  as  consistent  with  filling  and  cleaning.  All  wide-necked 
tubes  should  be  discarded.  The  behavior  of  the  closed  end  with  reference  to  the 

absorption  of  air  may  be  tested  by  adding  litmus- 
water  and  5  per  cent  grape-sugar  to  tlie  bouillon. 
On  steaming,  the  litmus  is  reduced.  If  there 
is  no  air  in  the  closed  end  the  litmus  remains 
reduced,  while  in  the  open  end  exposed  to  the  air 
it  soon  oxidizes  back  to  its  original  color. 

Other  things  to  be  observed  are : 

( j)   Growth  in  hydrogen. 

(<f)   Growth  in  carbon  dioxide. 

(5)  Growth  in  vacua,  various  degrees  of  ex- 
haustion. 

(6}  Growth  in  vacua,  remnant  of  oxygen  ab- 
sorbed by  the  mixture  of  caustic  potash  and  pyro- 
gallol  (same  as  pyrogallic  acid). 

(7)  Growth  in  nitrogen  (air  with  the  oxygen 
absorbed,  normal  air-pressure). 

The  hydrogen  and  carbon  dioxide,  which  are 
required  in  considerable  quantities,  may  be  gen- 
erated in  Kipp  gas-generators.  There  is  a  choice 
in  generators.  The  writer  has  not  found  any  kind 
which  is  entirely  satisfactory.  The  one  which  has 
given  the  least  trouble  is  shown  in  fig.  50.  The 
objection  to  this  generator  is  the  large  volume  of 
dead  acid  which  soon  accumulates  at  the  bottom. 
The  accumulation  of  dead  acid  is  entirely  obvi- 
ated in  the  de  Koninck  generator,  but  the  writer 
has  only  recently  obtained  this  apparatus  and  has 
not  yet  had  enough  experience  with  it  to  speak 
unqualifiedly.  It  furnishes  a  large  amount  of  gas 
and  its  generation  may  be  stopped  very  quickly, 
but  the  acid  chamber  is  inconveniently  bulky  (10 
liters)  and  in  case  of  breakage  a  destructive  flood 


Fig.  50.* 


would  be  poured  out  into  the  laboratory.     To  avoid  this  the  apparatus  should  be 
set  into  a  deep  enameled  iron  pan.     The  action  of  the  apparatus  depends  on  the  fact 


*FiG.  50.— Kipp  gas-generator  for  making  carbon  dioxide  or  hydrogen.  When  not  in  use  the 
pressure  of  the  gas  forces  the  acid  off  the  marble  or  zinc  (in  the  middle  compartment)  and  stops 
its  evolution.  Much  reduced. 


GAS-GENERATORS. 


55 


that  acid  on  which  zinc  has  reacted  has  a  greater  specific  gravity  than  unused  acid 
and  diffuses  downward  through  the  whole  fluid  when  it  is  forced  back  from  the 
zinc-chamber  into  the  top  of  the  acid-tank. 

Another  form  of  hydrogen  generator  is  shown  on  plate  7.  When  in  use  the 
lower  bulb  is  filled  with  acid  and  also  the  stem  of  the  upper  one.  This  gives  a 

sufficient  column  of  liquid  to  force  the  gas  through  the  five 
wash-bottles.  All  the  joints  should  be  coated  with  Darwin's 
wax-mixture,  set  together  firmly,  and  wired  in  place.  Exces- 
sive liberation  of  hydrogen  sulphide  is  avoided  by  standing 
the  generator  in  ice  water.  The  ruler  is  12  inches  long. 
The  same  style  of  apparatus  may  be  used  for  the  generation 
of  carbon  dioxide. 

These  gases  must,  of  course,  be  carefully  washed  to 
remove  accidental  poisonous  impurities,  by  passing  them 
through  wash-bottles  containing  various  solutions.  For  the 
carbon  dioxide,  which  is  usually  generated  from  c.p.  hydro- 
chloric acid,  diluted  with  twice  its  volume  of  boiled  water, 
and  marble  chips  (which  should  be  boiled  in  advance),  it  is 
sufficient  for  many  purposes  to  pass  it  through  strong  solu- 
tions of  sodium  hydrate  (10  per  cent),  potassium  permanga- 
nate (10  per  cent),  and  water,  arranged  in  the  order  indicated. 
Most  of  the  oxygen  may  be  removed  by  passing  through 
three  wash-bottles  containing  a  mixture  of  pyrogallol  and 
strong  caustic-potash  water  or  caustic-soda  water  (10  per 
cent).  When  in  use  the  stopcock  between  the  generator  and 
the  first  wash-bottle  must  not  be  cut  off,  otherwise  the  small 
amount  of  carbon  dioxide  in  the  wash-bottle  will  soon  be 
absorbed  by  the  soda  and  fluids  will  be  forced  over  (back- 
ward) from  the  other  bottles  by  inequalities  in  the  gas-pressure.  The  place  to  cut 
off  the  gas-flow  is  close  to  the  Novy  jar  or  other  receptacle. 

For  testing  the  purity  of  the  gas,  i.  e.,  its  freedom  from  air,  100  cc.  may  be 
drawn  off  into  a  Hempel  burette  (fig.  51),  equalized  with  the  air-pressure  and  run 
into  the  simple  Hempel  pipette  for  liquid  reagents  (fig.  52),  the  bulb  of  which  is 
filled  with  strong  potash  water  (2  water  +  i  potassium  hydroxide).  If  any  gas 
remains  after  thorough  exposure  to  the  potash,  it  may  be  measured  by  passing  it 
back  into  the  burette.  One  should  get  with  the  pipette  an  iron  stand  and  about 
2  yards  of  capillary  glass  tubing. 

The  scrap-zinc  used  for  generating  the  hydrogen  should  contain  some  lead,  but 
should  be  free  from  arsenic,  antimony,  and  phosphorus,  and  the  sulphuric  acid  should 
be  chemically  pure.  For  use  the  acid  is  diluted  largely  with  water  (1:9).  Hydro- 
gen generated  with  zinc,  especially  if  the  evolution  is  rapid  so  that  the  solution  is 
warmed,  contains  considerable  hydrogen  sulphide  and  may  contain  phosphureted 

*Fic.  51. — Hempel's  burettes  for  gas-analysis.  Height,  25  inches. 


Fig.  51.* 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


hydrogen  or  arseniureted  hydrogen ;  it  should  therefore  be  passed  not  too  rapidly 
through  the  following  solutions  in  the  order  indicated :  Saturated  solution  of  lead 
acetate,  5  per  cent  solution  of  silver  nitrate,  10  per  cent  potassium  permanganate, 
i o. per  cent  sodium  hydrate  containing  pyrogallol,  distilled  water.  When  ready 
for  use  the  purity  of  the  hydrogen  may  be  tested  by  burning  in  test-tubes  (mouth 

down  ,  and  also,  if  necessary,  by  the  ordinary 
methods  of  gas-analysis.  To  avoid  the  evolu- 
tion of  hydrogen  sulphide  the  generator  may  be 
plunged  into  a  jar  of  ice-water,  as  shown  in 
plate  7.  Special  care  must  be  taken  in  sealing 
jars  containing  hydrogen,  otherwise  it  will 
escape.  In  use,  the  gas  is  allowed  to  bubble 
slowly  through  the  fluids  into  the  culture- 
chamber,  a  large  well-clamped  Novy  jar,  the 
other  tubular  opening  of  which  is  connected 
air-tight  with  the  tube  of  the  vacuum  pipe. 
The  jar  is  first  pumped  out  and  the  hydrogen 
is  then  allowed  to  enter.  When  the  jar  is  full, 
the  glass  stopcock  nearest  it  (at  the  left  in  plate 
7)  is  turned,  and  then,  after  allowing  a  few 
minutes  for  diffusion,  the  mixture  of  air  and 
gas  is  pumped  out.  The  vacuum  cock  is  then 
turned  off  and  the  hydrogen  is  again  turned 
on  slowly.  This  process  is  repeated  five  or 
six  times,  the  gas  being  passed  into  the  jar 
very  slowly  the  last  two  times,  so  that  it  may 
be  washed  very  clean.  The  Novy  jar  is  then 
Fig.  52.*  sealed,  disconnected,  and  set  away  in  the  dark. 

The  gas  must,,  of  course,  enter  each  wash-bottle  through  the  long  stem.  It  is 
desirable  to  have  each  wash-bottle  two-thirds  full  of  fluid,  and  there  must  be  no  leaks 
in  any  part  of  the  apparatus.f  The  hydrogen  should  be  cut  off  before  each  exhaustion 
of  the  jar.  by  turning  the  stop-cock  nearest  the  jar.  The  cock  also  should  be  turned 
off  before  sealing  glass  tubes  with  flame  and  it  must,  of  course,  be  known  that  the 
gas  is  free  from  admixture  with  air,  otherwise  an  explosion  will  occur. 

It  is  easier  to  keep  air  out  of  gases  than  to  remove  it.  The  greatest  care 
should  therefore  be  taken  to  drive  it  out  of  a  culture  medium  before  it  is  inocu- 
lated. For  the  same  reason  gas  should  be  allowed  to  flow  for  some  time  before 
it  is  collected- so  as  to  displace  air  which  may  have  diffused  into  the  generator  and 
wash-bottles.  This  is  also  the  reason  why  the  water  which  is  used  to  dilute  the  acid 
and  the  marble  chips  should  be  boiled.  If  there  is  much  air  mixed  with  the  gas 
.t  is  riot  at  all  likely  that  a  single  wash-bottle  of  sodium  hydroxide  and  pyrogallol, 

*FiG.  52. — Hempel's  simple  pipette  for  liquid  reagents  used  in  gas-analysis.     Breadth  of  stand, 
7  inches. 

t  Consult  a  paper  by  Ewell,  Centralb.  f.  Bakt.,    2  Abt,  III  Bd.,  p.  188. 


PLATE  7. 


ft, 
f 


o 

K 


•8 

a 
rt 

•i 

s 

o 


5-   *! 
' 


a 

r 


f 

r 


ANAEROBIC    CULTURES. 


57 


or  even  two  or  three  in  series,  will  completely  remove  it,  since  the  bubbles  of 
gas  are  in  contact  with  the  fluid  only  at  their  surface  and  for  a  very  brief  time. 
Hydrogen  must  be  passed  through  5  wash-bottles  of  sodium  hydroxide  and  pyrogallol 
if  every  trace  of  oxygen  is  to  be  removed.  From  nitrogen  or  carbon  dioxide  the 
last  traces  of  oxygen  may  be  removed  by  passing  it  over  copper  filings  inclosed 
in  a  piece  of  gas-pipe  which  is  heated  red  hot  in  a  small  furnace  containing  about 
20  Bunsen  flames  in  series.  The  gas-pipe  may  be  0.75  inch  in  diameter  and  about 
3  feet  long,  plugged  at  the  ends  with  tight-fitting  rubber  stoppers,  the  middle  2  feet 
filled  with  the  copper  fragments.  The  gas  should  be  allowed  to  flow  only  in  rapid 
bubbles,  not  in  a  stream  (Dr.  Day). 

The  test-tube  cultures  may  be  placed  in  Novy  jars,  securely  waxed  (fig.  53), 

or  in  large,  thick-walled  test-tubes  made  impervious 
with  sealing  wax  (see  Sternberg,  Manual,  fig.  53 ; 
Text-Book,  fig.  53).  Media  designed  for  use  in  any 
of  these  gases  should  be  resteamed  immediately 
before  inoculation,  and  if  one  is  experimenting  with 
unknown  or  with  very  sensitive  anaerobes  the  boiled 
media  should  be  allowed  to  cool  in  an  atmosphere 
of  hydrogen.  Francis  Darwin's  wax-mixture  has 
been  found  useful  for  luting. 

When  large  Novy  jars  are  used  (fig.  54),  the 
thoroughly  waxed  gaskets  must  be  clamped  down 
securely  and  tested  for  leaks  by  preliminary  exhaus- 
tions. If  any  are  discovered,  additional  wax  must 
be  used  and  the  clamps  must  be  screwed  tighter.  To 
determine  whether  there  is  any  subsequent  entrance 
of  air  it  is  always  best  to  include  along  with  the 
cultures  one  or  more  tubes  containing  some  sub- 
stance which  is  reduced  in  the  absence  of  free  oxygen,  but  which  readily  oxidizes 
to  some  different  color  as  soon  as  traces  of  air  are  mixed  with  the  gas  in  the  jar. 
Methylene  blue  in  recently  steamed  bouillon  or  gelatin  with  5  per  cent  grape-sugar 
is  one  of  the  best  pigments  for  this  purpose.  In  the  absence  of  free  oxygen  it 
becomes  a  colorless  substance ;  with  the  entrance  of  traces  of  air  it  becomes  blue. 
Usually,  however,  the  fluid  or  solid  holds  on  to  a  trace  of  color  at  its  surface.  A 
solution  of  bilirubin  is  also  said  to  be  very  sensitive  to  free  oxygen  and  a  good  test 
for  its  presence. 

Some  care  is  necessary  in  order  to  avoid  erroneous  conclusions  when  pyro- 
gallol and  caustic  potash  are  used  to  absorb  the  oxygen.  The  vessel  must  not 
leak,  enough  of  the  mixture  must  be  used  to  absorb  all  the  oxygen,  and  the  action 
must  be  rapid  enough  so  that  the  oxygen  will  have  been  removed  completely  before 
visible  growth  of  the  organism  can  possibly  have  taken  place.  Neglect  of  these 


Fig.  53* 


*Fic.  53. — Novy  jar.     Small  size  (wide  mouth)  for  test-tube  cultures.     Only  those  with  mouths 
at  least  2*A  inches  wide  are  serviceable.     Height  to  mouth  of  jar,  7J4  'inches. 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


precautions  has  led  to  the  statement  that  certain  strict  aerobes  are  able  to  grow  on 
ordinary  media  in  the  absence  of  oxygen,  and  that  anaerobes  are  very  uncertain  in 
their  behavior  on  standard  media.  Old  pyrogallic  acid  should  be  avoided  and  some 
preliminary  experiments  should  be  made  as  to  the  rapidity  of  the  absorption  of  the 

oxygen  from  a  given  space  before  the 
organism  is  tested.  The  writer  found 
one  brand  of  pyrogallol  which  re- 
moved the  oxygen  from  a  small  space 
in  six  hours,  another  required  about 
eighteen  hours,  a  third  required  sev- 
eral days  (time  enough  for  a  strictly 
aerobic  organism  to  make  a  visible 
growth).  Leaks  may  be  detected  read- 
ily by  including  with  the  cultures 
a  fermentation-tube,  the  inclosed  arm 
filled  with  water  except  for  a  small 
bubble  of  airj  On  absorption  of  the 
oxygen  this  bubble  expands  to  a 
diameter  which  should  remain  con- 
stant if  the  jar  continues  air-tight. 

The  gas  remaining  in  receptacles 
from  which  the  oxygen  has  been 
removed  by  the  potash  -  pyrogallol 
method  is  not  pure  nitrogen,  but 
nitrogen  plus  a  variable  small  amount 
of  carbon  monoxide,  which  is  said 
to  be  most  abundant  when  the  oxy- 
gen is  absorbed  slowly.  This  small 
amount  of  CO  is  harmless  to  many 
bacteria,  but  the  writer  has  some 
reason  for  suspecting  that  it  is  inju- 


Fig.  54* 


rious  to  others,  even  if  it  does  not  entirely  inhibit  growth. 

The  writer  has  found  the  following  contrivance  (fig.  55)  a  very  simple  one  for 
testing  the  ability  of  organisms  to  grow  in  nitrogen  :  A  U-tube  of  thick,  clear  glass, 
with  arms  about  10  to  12  inches  long,  open  at  the  ends  and  having  a  uniform  inside 
diameter  of  about  i  inch,  serves  as  the  culture-chamber  and  gas-receptacle.  Two 
short,  rimless,  cotton-plugged  test-tubes  containing  the  media  to  be  tested  are  inocu- 
lated and  thrust  one  above  the  other  into  one  arm  of  the  U-tube,  into  which  is  then 


*Fic.  54. — Novy  jar  of  large  size  for  Petri  dishes  and  numerous  test-tube  cultures.  Clamped 
as  when  in  use.  Between  the  clamped  parts  is  a  rubber  gasket,  carefully  waxed  and  vaselined. 
Darwin's  wax>-mixture  is  advised.  The  writer  also  usually  wires  in  the  rwaxed  top  parts.  The  gas 
inflow  is  cut  off  by  twisting  the  uppermost  (horizontal)  ground-glass  stopper,  which  must  be  care- 
fully vaselined.  One-third  actual  size. 


ANAEROBIC    CULTURES. 


59 


, 


crowded  a  tight-fitting,  soft,  rubber  stopper.  This  end  is  finally  buried  for  an  inch 
or  so  in  a  small  beaker  of  glycerin  and  is  perfectly  air-tight.  A  rimless  test-tube 
about  5  inches  (13  cm.)  long  and  of  a  diameter  such  that  it  will  just  slip  easily  up  the 
other  arm  of  the  U-tube,  is  now  packed  by  means  of  a  pencil  or  glass  rod  with  8  or 

10  grams  of  pyrogallic  acid,  covered  quickly  with  25  cc. 
of  10  or  15  per  cent  caustic-potash  water,  and  slipped  up 
the  open  end  of  the  tube,  which  is  immediately  plunged 
into  a  dish  of  mercury  and  held  there  (under  a  shelf  )  until 
enough  of  the  oxygen  is  absorbed  so  that  it  will  stay 
down  of  its  own  weight.  The  exposure  should  be  made 
at  25°  or  30°  C.,  or  at  least  at  temperatures  considerably 
above  zero,  since  the  absorption  of  the  oxygen  is  slow  in 
cool  air. 

The  tube  containing  the  pyrogallic  acid  and  potash 
mixture  floats  on  the  mercury  and  rises,  of  course,  in  the 
arm  of  the  U-tube  as  the  oxygen  is  absorbed  and  the 
mercury  enters  it.  This  tube  must  not,  therefore,  be  too 
long  so  as  to  hit  against  the  curves  of  the  U-tube  before 
all  of  the  oxygen  has  been  absorbed  ;  otherwise  the  mer- 
cury will  pass  up  between  the  two  tubes  and  overflow 
into  the  mixture.  In  other  words,  several  centimeters 
must  be  allowed  for  the  rise  of  the  mercury. 

A  few  experiments  will  determine  how  much  of  the 
mixture  is  necessary  for  a  tube  of  a  given  bore  and  how 
long  it  takes  to  absorb  all  of  the  oxygen,  f  The  level 
of  the  mercury  in  the  open  end  with  all  the  oxygen 
absorbed  may  be  recorded  by  a  scratch  on  the  tube  as  a 
rough  guide  in  subsequent  work.  At  least  half  a  dozen 
of  these  tubes  will  be  found  useful.  They  may  be  made 
in  any  laboratory  or  may  be  procured  from  dealers  in 
glassware. 

In  the  use  of  carbon  dioxide,  especially  with  sensitive  organisms,  two  factors 
must  be  considered,  (i)  the  simple  exclusion  of  air,  as  in  case  of  hydrogen,  and 
(2)  the  change  in  the  reaction  of  the  medium  due  to  absorption  of  the  gas  (forma- 
tion of  carbonic  acid). 

*Fic.  55. — A  simple  device  for  growing  organisms  in  air  deprived  of  its  oxygen.  In  the  left 
arm  are  the  cultures ;  in  the  right  arm  is  a  test-tube  containing  a  mixture  of  pyrogallol  and  caustic- 
potash  water.  The  heaker  contains  mercury.  About  one-third  actual  size.  A  modification  of 
Ganong's  apparatus  for  study  of  germinating  seeds. 

fMace  states  that  I  gram  of  the  pyrogallol  and  10  cc.  of  the  10  per  cent  potash-water  are  suffi- 
cient for  each  100  cc.  of  air  space. 


Fig.  55.* 


6O  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

LUMINOSITY. 

Numerous  saprophytic  bacteria  are  luminous  under  certain  special  conditions. 
Luminosity  is  also  a  striking  characteristic  of  at  least  one  bacterial  animal  disease — 
the  white  disease,  or  sluggish  disease,  of  sand  fleas  (  Talorchestia  longicornis  and  T. 
megalophthalmid),  common  on  the  shores  of  France  and  ot  Massachusetts  at  Woods 
Hole.  Decaying  potatoes  and  other  vegetables  are  sometimes  luminous.  The 
question  of  luminosity  should  therefore  be  kept  in  mind  by  the  student  of  plant 
diseases,  although  no  luminous  species  are  known  to  live  in  plants.  Most  of 
these  interesting  luminous  bacteria  have  been  found  in  salt  water  or  near  it,  or  on 
the  flesh  of  quadrupeds  and  fish.  Gorham  has  been  able  to  grow  them  on  strictly 
synthetic  media.  The  most  recent  treatise  is  by  Hans  Molisch  (Leuchtende 
Pflanzen,  Jena,  Gustav  Fischer,  1904,  pp.  ix,  169,  with  2  plates  and  16  text  figures). 

Molisch  records  26  species  of  luminous  bacteria.  He  found  that  salt-water  fish 
and  the  flesh  of  cattle  exposed  in  the  markets  were  very  often  luminous — 48  per 
cent  of  70  samples  of  the  latter  and  nearly  all  the  former.  Of  horse  flesh  65  per  cent 
and  of  cattle  flesh  89  per  cent  became  luminous  on  putting  it  into  3  per  cent  solution 
of  sodium  chloride,  allowing  a  part  of  it  to  project  into  the  air.  Fresh-water  fish 
are  very  seldom  luminous.  Seedlings  exposed  to  Petri-dish  poured  plates  curved 
heliotropically  toward  the  light,  but  they  did  not  become  green.  Other  chlorides 
than  that  of  sodium  stimulate  growth  and  light-production,  e,  g.,  potassium, 
magnesium,  or  calcium  chloride.  Certain  non-chlorides,  such  as  potassium  iodide, 
potassium  sulfate,  and  magnesium  sulfate  have  the  same  action  (3  per  cent  or  less). 
Potassium  nitrate  was  also  active  on  B.  phosphoreum  but  not  on  B.  phologenus. 
Manganese  sulfate  stimulated  growth  very  noticeably  but  had  no  corresponding  effect 
on- the  luminosity,  which  was  weak.  The  spectrum  of  B.  phosphoreum  differs  from 
that  of  the  West  Indian  beetle,  Pyrophorus  noctilucus,  and  from  that  of  a  luminous 
fungus  known  as  mycelium  X.  No  biological  importance  is  attributed  to  the 
luminosity  which  is  ascribed  to  an  hypothetical  phologen.  It  is  an  oxidization 
phenomenon  which  can  take  place  only  in  the  presence  of  free  oxygen.  A  tem- 
perature of  30°  C.  for  forty-eight  hours  is  sufficient  to  kill  B.  phosphoreum  in  gelatin 
cultures.  The  minimum  temperature  for  this  organism  is  below  zero,  the  optimum 
is  about  16°  to  18°  C.,  and  the  maximum  is  28°  C.  The  bacteria  are  luminous 
from  minus  5°  to  plus  28°  C.  Light  production  is  most  intense  from  5°  to  20°  C. 

FERMENTATION   PRODUCTS. 

The  old  conception  of  fermentation  involves  an  evolution  of  gas  (fervere,  to 
boil),  but  the  term  is  now  used  with  a  wider  meaning.  Like  many  other  terms,  it  is 
difficult  to  use  it  always  logically.  In  general,  it  means  the  breaking  up  of  carbon 
compounds  into  simpler  substances,  either  by  the  direct  action  of  the  protoplasm  of 
the  organism  (hypothetical)  or  by  chemical  substances  (enzyms,  diastases)  secreted 
by  the  protoplasm.  Acids  and  alcohols  are  produced ;  gases  may  or  may  not  be 
evolved.  Other  volatile  products  are  also  produced,  e.  g.,  esters,  but  usually  only 
in  very  small  quantities.  Certain  of  the  bacterial  fermentations  are  of  large  com- 
mercial importance,  e.  g.,  the  acetic,  the  lactic.  The  breaking  up  of  albumen  and 


FERMENTATION    PRODUCTS.  6l 

other  complex  nitrogen  compounds,  i.  e.,  putrefaction,  is  also  sometimes  called  fer- 
mentation, and  at  present  there  is  really  no  very  sharp  line  to  be  drawn.  Consult 
Green  and  Duclaux  for  the  English  and  French  views  (Bibliog.,  XX).  The  student 
should  observe : 

(1)  Gases.  Amount,  rate  of  development,  kinds  (carbon  dioxide,  oxygen,  hydro- 
gen, nitrogen,  marsh  gas). 

(2)  Acids.     Volatile  and  non- volatile  (lactic,  acetic,  butyric,  etc.). 

(3)  Alcohols  (ethyl,  methyl,  butyl,  glycerin,  mannit,  etc.). 

(4)  Ethers  and  esters. 

(5)  Aldehyds,  sugars,  gums. 

(6)  Albumoses,  peptones,  amido-bodies. 

The  isolation  and  determination  of  the  amount  of  these  various  products 
belongs  to  the  province  of  the  chemist,  but  the  work  should  be  done  in  the  bacteri- 
ological laboratory  and  under  the  eye  of  the  biologist  if  all  sorts  of  errors,  due  to 
the  unsuspected  multiplication  of  intruding  organisms,  are  not  to  creep  in  and 
render  the  work  worthless.  Only  some  crude  determinations,  as  of  proportion  of 
the  various  gases  evolved,  may  be  made  by  the  bacteriologist  who  is  not  a  chemist. 
The  volume  of  gas  evolved  from  day  to  day  may  be  measured  in  fermentation-tubes 
(fig.  49).  Frost  has  devised  a  convenient  gasometer  for  roughly  estimating  it  (see  his 
Laboratory  Guide,  plate  I).  These  may  be  made  in  any  laboratory  out  of  cardboard. 

If  the  gas  is  carbon  dioxide  it  may  be  absorbed  by  shaking  with  10  per  cent 
NaOH.  To  do  this,  fill  the  bowl  (fig.  49)  even  full  of  the  strong  caustic-soda  water, 
place  the  thumb  or  forefinger  over  the  mouth  so  as  not  to  include  any  air,  invert  the 
tube  so  that  the  gas  shall  flow  into  the  bowl  and  come  into  contact  with  the  alkali, 
and  shake  vigorously  until  all  of  the  carbon  dioxide  is  absorbed.  Tilt  the  fluid 
back  into  the  open  end,  and  remove  the  finger  so  as  to  equalize  the  pressure.  If 
any  gas  remains  after  equalizing  the  air-pressure,  place  the  finger  over  the  mouth  of 
the  tube,  tilt  the  gas  into  the  bowl  and  apply  a  lighted  match  close  to  the  mouth  as 
the  finger  is  removed.  If  it  is  hydrogen  or  marsh  gas  it  will  explode  in  the  open 
end  of  the  tube  when  the  finger  is  removed  and  a  flame  applied.  If  it  is  nitrogen  it 
will  not  support  combustion  (see  Bibliog.,  XX,  especially  '90  Smith  and  '93  Smith). 

How  distinguish  marsh  gas  from  hydrogen  ? 

Organisms  easily  inhibited  by  their  own  acid  products  may  be  kept  alive  a 
much  longer  time  by  adding  a  little  calcium  carbonate  to  the  bouillon  or  agar. 

In  simple  tests  for  acids,  discard  bright  blue  litmus  paper,  which  is  very 
sensitive  to  carbonic  acid  (try  carbonated  water  on  it),  and  use  instead  a  good 
grade  of  reddish- violet  (neutral)  litmus  paper.  Such  paper  may  be  made  in  the 
laboratory  (the  best  way)  or  may  be  purchased  of  H.  Struers,  Copenhagen. 

ALKALIES  (AMMONIA,  AMINS,  CARBONATES  OF  THE  ALKALI  METALS). 

Determine  rapidity  of  formation.  Note  that  they  are  often  masked  by  the 
simultaneous  formation  of  acids.  Try  the  litmus  test  and  Nessler's  test  Do  not 
put  Nessler's  solution  into  the  culture  fluid,  but  expose  it  to  steam  from  the  culture. 
Observe  the  behavior  of  the  organism  when  grown  in  peptone  rosolic-acid  water 


62  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

with  just  enough  HC1  added  to  counteract  the  alkali  in  the  peptone,  and  in  neutral 
or  slightly  acid  peptone-water  or  sugar-free  bouillon  containing  acid  fuchsin.  On 
titration  of  acids  and  alkalies  see  Sutton  (Bibliography  of  General  Literature,  IV). 

REDUCING    POWERS. 

Determine  rapidity  of  reduction  of  litmus,  methylene  blue,  and  indigo  carmine 
in  various  fluids  and  solids  (with  and  without  grape-sugar).  Probably  all  bacteria 
can  reduce  litmus,  etc.,  but  as  the  rapidity  of  reduction  varies  greatly  in  different 
species  and  in  different  media,  it  is  desirable  to  make  comparative  tests.  Consult  a 
recent  paper  by  Albert  Maassen  ("Ueber  das  Reduktionsvermogen  der  Bakterien, 
und  ueber  reduzierende  Stoffe  in  pflanzlichen  und  tierischen  Zellen,"  Arb.  a.  d.  Kais. 
Gesuudheitsamte,  Bd.  XXI,  3  Heft,  1904,  pp.  377-384). 

HYDROGEN   SULPHIDE. 

This  gas  is  the  product  of  a  reduction.  From  what  media  and  under  what 
conditions  is  hydrogen  sulphide  given  off  with  browning  of  lead  acetate  paper  ? 
This  paper  is  readily  prepared  by  dipping  strips  of  white  filter  paper  into  a  strong 
solution  of  lead  acetate  in  distilled  water.  It  should  be  kept  in  a  tight  tin  box 
or  a  glass-stoppered  bottle.  Probably  most,  if  not  all,  bacteria  are  able  to  produce 
hydrogen  sulphide  in  nutrient  media  containing  readily  decomposable  sulphur  com- 
pounds. Is  an  enzyme  necessary  ?  When  an  organism  grays  potato  cylinders  in 
test-tubes,  why  is  no  hydrogen  sulphide  given  off?  The  student  should  read  papers 
by  Petri  and  Maassen  (Bibliog.,  XXVIII). 

MERCAPTAN  AND  OTHER  ODORS. 

We  need  an  odor  chart  to  go  along  with  our  color  charts.  If  we  could  have  a 
set  of  standard  substances  with  peculiar  smells  for  comparison  with  the  many 
odors  evolved  from  bacterial  cultures  it  would  certainly  be  a  great  convenience. 
The  difficulty  at  present  is  that  the  judgment  of  people  varies  greatly,  in  many 
instances,  as  to  what  the  smell  should  be  likened.  As  it  is,  the  bacteriologist  must 
do  the  best  he  can  to  define  these  penetrating  smells,  which  are  sometimes  very 
characteristic  of  particular  organisms.  Some  of  the  fishy  odors  are  due  to  am  ins. 
Mercaptan  is  a  very  vile-smelling  sulphur  compound. 

INDOL,  PHENOL,  LEUCIN,  TYROSIN,  ETC. 

The  production  of  indol  is  best  studied  in  peptonized  beef-broth  naturally  free 
from  sugar  or  which  has  been  deprived  of  its  muscle  sugar  by  growing  in  it  (for  a  few 
hours  only)  Bacillus  coli  (Theobald  Smith),  after  which  it  should  be  filtered  clear. 
If  B.  coli  or  B,  cloacae  will  not  produce  gas  in  beef-broth  in  the  closed  end  of 
fermentation-tubes,  it  is  free  from  sugar  and  suitable  for  this  use.  Many  organisms 
give  the  indol  reaction  in  Uschinsky's  solution  to  which  peptone  has  been  added. 
The  writer  has  never  been  able  to  obtain  the  indol  reaction  in  any  culture  medium 
which  did  not  contain  peptone  (using  this  word  in  the  commercial  sense. )  Cultures 
which  do  not  show  the  red  reaction  with  sodium  nitrite  (0.02  per  cent  solution)  and 
sulphuric  acid  at  room  temperature  will  frequently  do  so  when  put  into  hot  water 


INDOL,    PHENOL,    ETC. 


for  five  minutes  (70°  to  80°  C.).  The  browning  of  media  due  to  excess  of  sodium 
nitrite  must  not  be  mistaken  for  this  pink  or  red  reaction.  Uninoculated  tubes 
should  be  included  in  the  test,  which  may  be  made  on  the  second  and  tenth  day. 

For  methods  of  determining  phenol  see  Lewandowski  in  Deutsche  Med. 
Wochenschrift,  1890,  p.  1186,  and  Chester's  Manual,  p.  33.  Schmidt  (Bd.  II, 
p.  1008)  gives  the  following  as  a  qualitative  reaction  for  tyrosin  :  Dissolve  by 
boiling  in  water  and  add  a  solution  of  mercuric  nitrate.  The  red  reaction  is 
sharper  if  a  little  fuming  nitric  acid  diluted  in  water  is  added.  Try  also  the  violet 
reaction  with  neutral  iron  chloride.*  Leucin  crystallizes  in  white  soft  scales. 

REDUCTION   OF   NITRATES   (AND   MORE   COMPLEX   NITROGEN   COMPOUNDS)   TO 
NITRITES,   TO   AMMONIA,    AND  TO   FREE   NITROGEN. 

For  the  pathologist  the  iodine-starch  reaction  is  the  most  satisfactory  test  for 
nitrites,  because  it  is  not  superlatively  sensitive  and  consequently  does  not  indicate 

traces  of  nitrite  absorbed  from  the  air.  It  is  made  as 
follows :  Twenty-five  cubic  centimeters  of  distilled 
water  are  added  to  one-half  gram  (more  or  less)  of 
pure  potato  starch  and  the  fluid  boiled.  One  cubic 
centimeter  or  more  of  this  starch-water  and  i  cc.  of 
freshly  prepared  potassium-iodide  water  (i  :  250)  are 
now  put  into  the  culture  fluid,  to  which  is  then  added 
a  few  drops  of  strong  sulphuric-acid  water  (2  :  i).  If 
any  appreciable  quantity  of  nitrite  is  present  the  culture 
immediately  becomes  blue-black  from  the  liberation  of 
free  iodine,  which  acts  upon  the  starch.  Old  potassium 
iodide  water  should  never  be  used  without  first  testing 
carefully,  as  it  usually  contains  some  free  iodine.  It 
is  always  best  to  first  make  a  trial  test  without  the 
bacteria.  Commercial  starch  frequently  contains  prod- 
ucts of  bacterial  decomposition  and  starch  prepared 
aseptically  should  be  substituted. 

At  least  one-third  of  the  organisms  which  have 
fallen  under  the  writer's  observation   in  recent  years 
give  the  nitrite  reaction  when  grown    in  peptonized 
beef-bouillon  containing  potassium  nitrate. 

*Mann  (p.  323)  gives  the  following  as  a  specific  tyrosin  reaction:  Deniges  has  recommended 
the  well-known  phenol  aldehyde  reaction  for  the  detection  of  tyrosin.  Nasse,  in  repeating  Deniges' 
observations,  has  found  the  following  to  be  a  very  delicate  test  for  tyrosin,  as  neither  proteids 
nor  peptones  give  the  color-reaction.  Proceed  thus:  Add  a  few  drops  of  formol  solution  to  con- 
centrated sulphuric  acid,  when,  on  warming  with  tyrosin,  a  brown-red  color  is  obtained,  which,  on 
addition  of  acetic  acid,  becomes  green. 

fFic.  56.— Bacterium  syringae  (van  Hall).  Nitrate  bouillon  cultures  5  days  old,  to  each  of 
which  has  been  added  boiled  starch  water,  potassium  iodide  water,  and  sulphuric  acid.  In  tube  a 
the  potassium  nitrate  was  reduced  to  the  nitrite,  and  on  addition  of  the  reagents  free  iodine  was 
liberated,  and  the  starch  blued.  In  the  other  no  nitrite  had  formed,  no  iodine  was  liberated,  and  the 
starch  remained  colorless.  For  discrepancy  see  text 


Fig.  56.t 


64  BACTERIA   IN    RELATION   TO    PLANT   DISEASES. 

Fig.  56  shows  how  differently  quite  similar-looking  cultures  may  react  when 
submitted  to  this  test.  Both  of  these  organisms  were  received  from  van  Hall  under 
the  name  of  Pseudomonas  syringe,  a  being  van  Hall's  own  isolation  and  b  being 
supposedly  a  subculture  from  Beyerinck's  isolation.  Neither  one  would  produce 
any  blight  in  lilac  shoots. 

There  is  no  simple  way  known  to  the  writer  of  distinguishing  ammonia  from 
the  amins,  as  both  react  to  Nessler's  reagent  Nitrogen  may  be  distinguished  from 
the  other  gases  of  fermentation  by  the  fact  that  it  is  not  absorbed  by  sodium  or 
potassium  hydroxide  and  will  not  burn  or  support  combustion.  This  gas  is  produced 
readily  from  nitrates  by  a  number  of  green-fluorescent  organisms  (dung-destroyers) 
but  not  by  all  of  them. 

FIXATION  OF  FREE  NITROGEN  AND  THE  OXIDATION  OF  AMMONIA  AND  AMMONIUM 

SALTS  TO  NITRITES  AND  NITRATES. 

These  processes  are  probably  common  enough  to  organisms  of  the  soil,  many 
of  which  have  not  been  investigated,  but  they  are  not  known  to  be  brought  about  by 
plant  parasites  exclusive  of  the  root-tubercle  bacilli  of  the  Leguminosae,  which  some 
believe  to  be  parasites  (see  Peirce).*  They  are  believed  to  be  of  rare  occurrence  in 
bacteria  which  grow  well  on  ordinary  culture  media. 

The  nodules  on  roots  of  plants  will  hereafter  be  considered  more  fully.  The 
reader  should  consult  a  paper  by  Geo.  T.  Moore  on  "  Soil  Inoculation  for  Legumes," 
Bureau  of  Plant  Industry,  United  States  Department  of  Agriculture,  Bull.  71,  Jan- 
uary 23, 1905  ;  and  one  by  Maria  Dawson,  "Further  Observations  on  the  Nature  and 
Functions  of  the  Nodules  of  Leguminous  Plants,"  Philosophical  Transactions 
Royal  Society  of  London,  Series  B,  Vol.  CXCIII,  pp.  51-67,  1900,  with  2  plates. 

ASSIMILATION   OF   CARBON    DIOXIDE. 

Some  soil  organisms  are  believed  to  obtain  their  carbon  directly  from  carbon 
dioxide,  and  would  thus  be  exceptions  to  the  law  that  all  non-chlorophyllous  plants 
must  obtain  their  carbon  from  organic  substances.  This  supposition,  while  probably 
true,  has  not,  we  believe,  been  established  satisfactorily.  Its  elucidation  offers  a 
most  interesting  line  of  research  (see  Bibliog.,  XXVI.) 

PIGMENTS. 

Bacterial  growths  are  often  bright  colored,  and  an  examination  of  the  pigments 
should  form  part  of  one's  study  of  an  organism.  They  may  be  considered  as  follows  : 

(1)  Under  what  conditions  formed  ?      Can  they  be  eliminated  by  growing  the 
organisms  in  the  dark  or  under  unfavorable  conditions,  e.g.,  near  the  maximum  or 
minimum  temperature  ?     Bacillus prodigiosus  is  a  favorable  organism  for  experiment. 

(2)  In  what  soluble  (water,  hydrogen-peroxide  in  water,  ethyl  alcohol,  methyl 
alcohol,  glycerin,  acetic  ether,  petroleum  ether,  sulphuric  ether,  acetone,  chloroform, 
turpentine,  benzine,  benzole,  xylol,  toluol,  carbon  bisulphide,  etc.)?     The  pigment 
should  be  tested  in  as  many  solvents  as  possible. 

*Peirce,  George  James.  The  Root-tubercles  of  Bur  Clover  (Medicago  denticulata  Willd.)  and 
of  Some  Other  Leguminous  Plants.  Proc.  Calif.  Acad.  Sci.,  3d  series,  Botany,  Vol.  II,  No.  10,  San 
Francisco,  Cal.,  June  21,  1902,  pp.  295-328,  with  i  plate. 


PIGMENTS.  65 

(3)  How  are  they  acted  on  by  acids,  alkalies,  and  other  reagents  ? 

(4)  Of  what  use  are  they  to  the  organism  ?     Are  they  oxidation-products  ? 
Examine  spectroscopically,  if  possible. 

On  the  addition  of  acids  or  alkalies,  a  bacterial  pigment  may  remain  unchanged, 
may  be  changed  into  some  different  color,  may  be  destroyed,  or  may  be  converted 
into  some  colorless  compound  which  will  regain  its  original  color  on  changing  back 
the  reaction.  The  yellow  pigment  of  several  species  of  Bacterium  (Pseudomonas) 
remains  unchanged  in  the  presence  of  acids  and  alkalies.  The  blood-red  color  of 
Bacillus  prodigiosus  becomes  carmine  in  the  presence  of  certain  acids  and  yellowish- 
brown  in  the  presence  of  certain  alkalies.  The  blue  color  of  Bacterium  syncyaneum 
is  said  to  be  produced  only  in  acid  milk.  The  beautiful  green  fluorescence  of  Bac- 
terium pericarditidis  (Bacillus  pyocyaneus  pericarditidis~),  and  probably  of  all  this 
group  of  bacteria,  is  produced  only  in  alkaline  media.  According  to  Jordan  two 
pigments  are  normally  produced  by  many  green-fluorescent  organisms.  The  blue 
pigment  pyocyanin  is  visible  by  gaslight  and  is  soluble  in  chloroform.  The  green- 
fluorescent  pigment  is  insoluble  in  chloroform  and  yellowish  by  gaslight.  By 
this  latter  test  the  two  can  be  distinguished  when  mixed.  Soluble  phosphates  and 
sulphates  are  necessary  for  the  production  of  green  fluorescence.  The  ability  to 
produce  pyocyanin  is  easily  lost.  Its  production  in  the  culture-medium,  unlike  that 
of  the  fluorescine,  is  not  dependent  on  the  presence  of  phosphates  or  sulfates. 
Pyocyanin  turns  red  with  acids,  fluorescine  becomes  colorless ;  both  return  to  their 
original  color  on  adding  alkali  sufficient  to  change  the  reaction.  "  Asparagin, 
ammonium  succinate,  ammonium  lactate,  and  ammonium  citrate  all  proved  suitable 
for  the  development  of  the  fluorescent  pigment."  The  yellow  and  black  pigments 
are  the  result  of  oxidations.  (See  papers  by  Gessard,  Thumm,  and  Jordan,  Bibliog., 
XXIII). 

The  pigments  of  bacteria  range  from  one  end  of  the  spectrum  to  the  other. 
Thus  we  have  various  shades  of  black,  brown,  violet,  indigo,  blue,  green,  yellow, 
orange,  and  red.  Many  bacteria  produce  no  pigment,  i.  e.,  are  white  when  seen  in 
mass.  Others  produce  several  distinct  pigments.  Many  of  the  plant  parasites  are 
yellow,  e.  g.,  Bacterium  campestre,  Bact.  phaseoli,  Bad.  hyacinthi,  Bact.  Stewarti, 
Bad.  juglandis.  Some  of  these  yellow  organisms  stain  the  host-plant  and  certain 
nutrient  substrata  a  deep  brown.  Other  plant  parasites  are  white  but  also  stain  the 
host  and  certain  substrata  brown,  e.g,  Bacterium  solanacearum,  Bacillus  carotovorus, 
B.  aroidece.  Others  are  pure  white  and  are  apparently  destitute  of  any  pigment- 
producing  powers,  e.g.,  Bacillus  amylovorus,  B.  tracheiphilus.  Very  many  bacteria 
when  grown  on  cooked  potato  produce  a  gray  stain  in  this  substratum,  especially  in 
that  part  freely  exposed  to  the  air,  i.  e.,  out  of  the  water. 

Some  other  color  changes  in  the  host  should  be  mentioned.  Various  brown 
and  red  stains  visible  in  certain  plants  when  attacked  by  bacteria  are  not  attributable 
directly  to  the  presence  of  the  microorganisms  in  the  tissues.  These  are  oxidation 
phenomena  likely  to  occur  when  the  plants  are  wounded  or  destroyed  by  any  agent 
whatsoever.  A  few  illustrations  will  make  my  meaning  clear.  When  the  limbs  of 
pear  trees  are  destroyed  by  blight  the  foliage  becomes  black,  but  this  blackening 


66  BACTERIA   IN    RELATION   TO    PLANT   DISEASES. 

also  occurs  frequently  when  the  flowers,  green  fruits,  or  foliage  are  killed  by  other 
causes.  In  the  leaves  of  Amaryllis  atamasco  the  writer  obtained  red  stripes  by 
injecting  the  yellow  Bacterium  hyacinthi,  but  no  bacterial  disease  followed,  and  the 
same  plant  reddens  when  bruised.  Broomcorn  shows  conspicuous  red  blotches 
when  attacked  by  the  broomcorn  organism,  but  the  parasite  itself  does  not  produce 
a  red  pigment,  while  the  plant  reddens  easily  as  the  result  of  aphis-punctures  or 
wounds  of  any  sort.  Sugar-cane  attacked  by  Bacterium  vascularum  shows  a  con- 
spicuous red  stain  in  the  bundles,  but  other  causes,  such  as  the  gnawings  of  an  insect 
or  the  presence  of  a  fungus,  may  lead  to  a  similar  stain,  while  the  bacterium  itself 
does  not  produce  any  red  pigment. 

CRYSTALS. 

Determine  the  nature  of  the  crystals  observed  in  the  various  media.  Many  of 
these  are  double  ammonium  salts;  others  result  from  the  action  of.trypsin  on  pro- 
teids.  Crystals  which  are  not  due  to  the  drying  out  of  the  media  are  common 

phenomena  in  old  ciiltures  of  many  sorts,  especially  if 
the  media  were  not  originally  saturated  with  alkali 
(soda  or  potash).  Fig.  57  shows  two  types  of  crystals 
formed  in  +15  nutrient  agar  by  two  green-fluores- 
cent organisms  received  from  van  Hall  as  Pseudomonas 
syringte,  and  a  third  type  produced  by  the  olive 
tubercle  organism. 

QUESTION   OK   EXISTENCE   OF   ENZYMES. 

The  enzymes  of  English  writers  are  the  diastases 
of  Duclaux.  They  are  chemical  substances,  the  exact 
composition  of  which  has  not  been  determined.    They 
Fig.  57.*  may  be  regarded  as  the  working  tools  of  protoplasm. 

The  following  are  some  of  the  best  known  kinds  : 

(i.)  Diastasic  (starch-destroying).  (5.)  Lab  or  rennet  (casein-forming). 

(2.)  Inverting  (sugar-splitting).  (6.)  Lipase  (fat-splitting). 

(3.)  Cytohydrolytic  (cellulose-dissolving.)  (7.)  Pectic  (pectin-splitting). 

(4.)  Proteolytic  (peptonizing).  (8.)  Oxidases  (oxidizing). 

Trypsin  is  common.  Pepsin  is  not  known  to  be  produced  by  bacteria  and 
should  be  searched  for. 

Many  bacteria  invert  cane-sugar,  but  invertase  is  believed  to  be  rare.  This, 
however,  may  be  an  ill-founded  conclusion.  The  experiments  of  various  animal 
physiologists  have  shown  that  when  cane-sugar  is  injected  into  the  blood-stream  it 
is  excreted  unchanged,  and  according  to  Julius  Sachs  cane-sugar,  inulin,  etc.,  must 

*FiG.  57. — Crystals  formed  in  cultures  of  Bacterium  syringae  (van  Hall).  I.  From  tube  II, 
Aug.  14  (agar  stock  693),  from  van  Hall's  II,  i.e., his  own  isolation  corresponding  to  a,  fig.  56. 
2.  From  tube  I,  Aug.  14  (stock  693),  from  van  Hall's  I,  which  is  from  Beyerinck's  old  isolation  (see 
b,  fig.  56)  X  3.  Nos.  i  and  2  drawn  Aug.  30, 1902.  3.  Crystals  formed  on  slant  litmus-lactose  agar 
which  was  inoculated  with  the  organism  causing  olive-knot.  About  one-half  inch  of  slant  in  middle 
part  of  culture  i  month  old,  i.  e.,  made  January  20,  1904;  drawn  February  17-19.  X  3-  Tempera- 
ture during  growth,  20°  to  25°  C. 


ENZYMES.  67 

first  be  reduced  to  glucose  (grape-sugar),  before  they  can  be  used  as  food  by  plants. 
When  no  invertase  has  been  detected  the  general  hypothesis  has  been  that  this 
inversion  was  due  to  the  direct  action  of  the  protoplasm,  but  the  recent  isolation  by 
Buchner  and  others  of  an  invertase  (Zymase)  from  yeast,  in  which  it  was  long  believed 
that  none  existed,  once  more  emphasizes  the  uncertainty  of  negative  conclusions. 
Diastase  is  common.  Is  there  more  than  one  kind,  i.  e.,  a  sort  which  can  only 
convert  the  starch  into  amylodextrin  and  another  which  converts  it  into  maltose 
and  dextrine  ?  In  many  cases,  when  the  organism  is  grown  on  potato,  the  con- 
version is  carried  only  a  little  way  and  stops,  there  being  always  a  copious  purple 
or  red-purple  reaction  with  iodine.  In  other  cases,  e.g.,  when  Bacterium  campestre 
is  grown  on  potato,  the  starch  conversion  is  so  complete  that  after  a  few  weeks  there 
is  little  or  no  color  reaction  when  the  potato-cylinder  is  mashed  up  and  iodine  water 

added.     What  makes  this  difference? 

A  substance  capable  of  dissolving  the  middle 
lamella  appears  to  be  common  to  all  bacterial  plant 
parasites  and  a  true  cytase  presumably  occurs,  but 
much  additional  study  is  necessary.  Probably 
several  enzymes  are  confused  under  this  name, 
just  as  several  chemically  different  substances  are 
still  called  "cellulose."  The  substance  which 
dissolves  the  middle  lamella  in  some  cases  is  prob- 
ably ammonium  oxalate.  The  writer  has  not  been 
able  to  dissolve  it  by  means  of  pure  oxalic  acid, 
but  that  of  turnips  softens  in  ammonium  oxalate. 
The  lab  or  rennet  ferment  is  rather  common. 
Its  action  should  not  be  confused  with  the  curdling 
of  milk  due  to  the  formation  of  acids.  Tests  may 
be  made  in  litmus  milk.  Is  there  more  than  one 
kind  of  such  ferment?  Some  organisms  coagu- 
late the  milk  promptly  into  a  solid  mass  which 
finally  shrinks,  extruding  whey.  Others  cause  the 
Fig-  58,*  casein  to  separate  out  of  the  fluid  very  slowly  as  a 

multitude  of  separate  particles  which  only  become  compacted  very  slowly. 

The  writer  has  not  met  with  the  oxidizing  enzymes,  unless  the  substance  in 
bacterial  cultures  which  causes  rapid  evolution  of  oxygen  from  hydrogen  peroxide  is 
such  an  enzyme,  as  Dr.  Loew  maintains  (Bibliog.,  XLV).  Many  other  enzymes 
undoubtedly  occur  and  play  their  part.  The  student  should  search  for  emulsin, 
lipase,  lactase,  maltase  (glucase),  etc. 

All  known  enzymes  when  freely  exposed  to  steam  heat  are  destroyed  at  tempera- 
tures considerably  under  100°  C.  They  are  less  sensitive  to  heat  than  the  bacteria 
themselves,  but  are  destroyed  by  a  few  minutes  exposure  to  temperatures  15°  to  30°  C. 
(moist  heat)  above  the  thermal  death-point  of  the  organisms  which  have  produced 

*Fic.  58. — Thick-walled  Kitasato  flask  for  filtration   or   evaporation   in  vacua,  etc.     Much   re- 
duced. 


68 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


Fig.59.f 


them.*  Some  of  them  are  very  sen- 
sitive to  the  presence  of  acids,  alka- 
lies, strong  alcohol,  or  antiseptics,  or 
their  action  is  inhibited  by  the  pres- 
ence of  other  enzymes  or  of  products 
of  enzymic  fermentation  in  excess,  or 
by  the  absence  of  some  combining 
substance,  such  as  lime  or  some  weak 
acid.  Some  do  not  pass  readily 
through  the  Chamberland  filter  or 
through  filter  papers.  Some  are 
destroyed  at  lower  temperatures  after 
precipitation.  Some  are  not  pro- 
duced except  in  presence  of  the  sub- 
stance which  they  can  decompose, 
but  this  is  not  true  of  all.  Usually 
an  organism  produces  more  than 
one  ferment  and  some  bacteria  are 
known  to  produce  five  or  six.  Bac- 
terium campestre  produces  at  least 
three  and  probably  four,  viz,  diasta- 
sic,  cytohydrolytic,  proteolytic,  and 
rennet.  It  also  inverts  cane-sugar, 
but  it  is  not  yet  known  whether  this 
change  is  accomplished  by  means  of 
an  invertase.  On  enzymes  derived 
from  bacterial  soft-rot  organisms  the 
reader  should  consult  recent  papers 
by  Jones  (Centralb.  f.  Bakt,  2  Abt.,  and 
Vermont  Exp.  Sta.  Rep.).  Levy  has 
published  an  interesting  paper  on 
"  Some  physical  properties  of  en- 
zymes" (The  Jotir.  Infect.  Diseases, 
Vol.  II,  1905,  pp.  1-48). 

For  concentrating  fluids  in  vacuo 
at  low  temperatures  (50°  to  60°  C.) 
the  thick-walled  Kitasato  flask  shown 


*The  same  amount  of  dry  heat  does  not  affect  them,  and  Loeffler  has  recently  advised  exposure 
of  thoroughly  air-dried  tissues  and  cultures  to  150°  C.,  dry  heat,  as  an  easy  way  of  eliminating  the 
bacteria  prior  to  grinding  and  extraction  of  the  uninjured  enzymes  and  other  soluble  products. 
Non-sporiferous  bacteria  may  be  heated  at  120°  C.  for  2  to  3  hours.  Tissues  and  sporiferous  bacteria 
should  be  heated  at  150°  C.  for  one-half  hour.  (Deutsche  Med.  Wochenschrift,  Dec.  22,  1904.) 

fFic.  59.— ^Burettes  used  by  the  writer  for  titrating  culture  media.  Twentieth-normal  sodium 
hydrate  is  used  to  determine  the  acidity, 'and  the  medium  is  finally  brought  to  the  desired  alkalinity 
with  quadruple-normal  sodium  hydrate.  The  fluid  is  boiled  and  titrated  hot,  using  phenolphthalein 
as  the  indicator.  The  burettes  should  be  graduated  to  tenths  of  a  cubic  centimeter  and  should  hold 
50  cc.  Alkali  should  not  be  allowed  to  stand  in  them. 


EVAPORATION    AT    LOW    TEMPERATURES. 


iii  fig.  58  is  very  convenient.  The  side  tube  is  attached  to  the  suction-pipe  of  an 
air-pump  and  into  the  neck  is  thrust  a  rubber  stopper  carrying  a  thermometer  and 
a  U-shaped  glass  tube  of  small  bore,  the  outer  arm  (36  inches  long)  ending  in  a 
beaker  of  mercury.  Heat  may  be  applied  by  means  of  a  water-bath.  By  substitut- 
ing a  funnel  for  the  thermometer  the  same  device  may  be  used  to  hasten  the  filtration 
of  thick  liquids,  hard-pointed  filter  papers  being  employed. 

SENSITIVENESS  TO   PLANT   ACIDS. 

The  tests  should  be  made  with  malic,  citric,  lactic,  oxalic,  and  tartaric  acids 
added  to  neutral  beef-broth,  peptone-water,  or  plant-broths,  or  to  synthetic  media 

(see  Am.  Nat.,  1899, p.  208).     It  is  best  to  titrate  with—  or  —  solutions,  to  acidify 

2N       4.N 
with  -  —  or  —  -  solutions,  and  to  reckon  the  acidity  in  cubic  centimeters  of  normal 

N 

solution  (— )  required  per  liter  of  medium.  If  pre- 
ferred, it  may  be  calculated  on  100  cc.  portions  and 
expressed  in  per  cents,  but  there  is  no  advantage  in  this, 
and  it  has  the  disadvantage  of  introducing  fractions. 

SENSITIVENESS  TO  ALKALIES  (POTASSIUM  OR  SODIUM 
HYDRATE). 

Determine  in  each  case  the  optimum  reaction  of  the 
medium  for  growth.  For  the  majority  of  bacteria  this 
is  said  to  lie  between  +10  and +  15  of  Fuller's  scale.f 
The  best  neutral  litmus  paper  should  be  used  freely,  but 
acid  and  alkaline  media  should  be  titrated  with  phenol- 

N        N 

phthalein   and  —  or  —   solutions.     In  some  media — 
10       20 

e.g.,  gelatin,  juices  of  various  plants — the  end-reaction 
with  phenolphthalein  and  caustic  soda  is  not  very  sharp. 
In  these  cases  the  titration  should  be  stopped  at  the  first 
trace  of  change  of  color.  If  one  adds  alkali  until  the 
fluid  is  decidedly  red,  then  a  distinct  statement  to  that 
effect  should  be  made,  since  otherwise  no  comparisons  of 
any  value  can  be  made.  All  of  the  writer's  +  and  — refer- 
ences to  media  are  based  on  a  reaction  stopped  at  the 
first  distinct  trace  of  pink  color.  As  much  again  alkali 
must  sometimes  be  added  to  obtain  a  deep-red  color. 

*Fic.  60. — Stock  bottle  of  —  sodium  hydrate  solution.  The  small  bottle  at  the  right  holds  con- 
centrated potash  liquor  to  remove  the  carbon  dioxide  from  the  air  which  enters  the  bottle.  About 
one-fourth  actual  size. 

fThe  plus  and  minus  on  Fuller's  scale  denotes,  respectively,  acid  and  alkaline  media.  The  +  ID, 
for  example,  means  that  exactly  10  cubic  centimeters  of  normal  alkali  must  be  added  to  a  liter  of 
the  culture  medium  to  render  it  exactly  neutral  to  phenolphthalein,  and,  correspondingly,  — 10  means 
that  the  fluid  is  alkaline  to  phenolphthalein  and  that  10  cc.  of  normal  acid  would  need  to  be  added  to 
bring  I  liter  back  to  the  neutral  point.  The  student  should  not  confuse  the  litmus  neutral  point  and 
the  phenolphthalein  neutral  point,  as  they  are  about  23°  apart,  e.  g.,  + 10  of  Fuller's  scale  (acid  side) 
is  distinctly  alkaline  to  litmus.  (Consult  '95,  Fuller,  Bibliog.,  XVI.) 


70  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

The  writer  has  used  the  foregoing  method  of  determining  the  reaction  of  culture 
media  for  several  years  and  has,  in  general,  found  it  exceedingly  exact  and  valuable, 
but  it  does  not  appear  to  be  well  adapted  for  determining  the  amount  of  alkali 
(ammonia  and  amins)  produced  by  bacteria  in  culture  media  (see  Button,  Bibliog., 
IV).  The  apparatus  required  to  make  these  titrations  is  shown  in  figs.  59  and  60. 

Some  experiments  recently  made  by  the  writer  with  Bacillus  tracheiphilus  in 
peptonized  beef-bouillons  of  varying  degrees  of  acidity  (acid  of  beef-juice)  and  alka- 
linity seem  to  show  that  toleration  of  sodium  hydrate  can  be  considerably  increased  by 
inoculating  each  time  from  alkaline  bouillons  rather  than  from  acid  ones.  Taken 
from  +  20  bouillon  (descended  from  +  20  bouillon)  this  organism  would  cloud  the 
same  bouillon  only  down  to  o;  taken  from  o  or  — 5  bouillons  (descended  from  — 2.7 
bouillon)  it  would  cloud  the  same  bouillon  down  to  — 10  and  probably  farther,  but 
not  to  — 20.  Bouillon  containing  various  amounts  ofc.  p.  sodium  chloride  behaved 
in  the  same  way.  The  organism  would  tolerate  the  largest  amount  of  salt  (1.5  to 
2  per  cent)  when  first  grown  in  an  alkaline  bouillon.  When  inoculated  from  a  +20 
bouillon  the  organism  finally  grew  in  i  per  cent  salt  bouillon,  but  only  after  a 
decided  retardation,  and  would  not  grow  at  all  in  + 1 5  peptonized  beef-bouillon 
containing  1.5  per  cent  sodium  chloride. 

Bacteria  vary  greatly  in  their  toleration  of  acids  and  alkalies,  the  range  of  growth 
being  from  minus  100  (or  more)  of  Fuller's  scale  to  phis  100  (or  more).  The  limits 
of  growth  are  not  known,  but  it  is  probable  that  the  extremes  of  toleration  in 
particular  aberrant  species  is  much  greater  than  that  here  given,  e.  g.,  on  the  acid 
side  in  sulphuric  acid  and  vinegar  bacteria,  and  on  the  alkaline  side  in  case  of  those 
organisms  which  are  able  to  grow  in  the  lime-vats  of  tanning  establishments 
and  in  alkaline  springs.  Lehmann  &  Neumann  ('96,  Bibliog.,  Ill),  state  that 
they  have  found  bacteria  that  will  endure  100  cc.  of  normal  acid  per  liter  of  fluid 
culture  media,  i.  £.,  equal  to  about  i  per  cent  sulphuric  acid.  Some  species  are 
indifferent  to  a  considerable  degree,  having  a  wide  range  of  growth  either  side  of 
the  (phenolphthalein)  neutral  line  ;  others  prefer  alkaline  media ;  others  acid  media. 
Many  are  extremely  sensitive  to  their  own  acid  products  (acetic,  lactic,  butyric,  etc., 
acids).  Not  a  few  are  differently  affected  by  different  acids  and  alkalies.  Every 
new  organism  presents  a  whole  series  of  special  problems. 

EFFECT  OF  DESICCATION. 

Drops  of  fluid  cultures  or  small  masses  of  gelatin  or  agar  cultures  are  spread  on 
small  ("4 -inch)  clean,  sterile  cover-glasses,  in  covered  sterile  Petri  dishes,  and  are 
set  away  in  the  dark,  in  dry  air  (a  dry  room).  The  test  is  finally  made  by  seizing 
one  of  these  covers  with  a  pair  of  sterile  forceps  and  dropping  it  into  a  tube  of  sterile 
bouillon  or  other  medium  of  a  stock  previously  determined  to  be  exactly  adapted  to 
the  growth  of  the  organism,  /.  e.,  one  which  does  not  exert  upon  it  any  retarding 
influence.  Occasionally  a  tube  will  become  contaminated,  but  enough  must  be 
inoculated  so  that  this  will  not  affect  the  final  result  (20  at  one  time  is  not  too 
many).  Fluid  cultures  are  preferred.  Solid  cultures  do  not  give  strictly  compar- 
able results. 


EFFECT   OF    DESICCATION.  Jl 

Organisms  believed  to  be  non-sporiferotts  show  great  differences,  some  being 
killed  by  an  exposure  of  a  few  minutes  or  a  few  hours,  while  others  remain  alive  for 
many  weeks.  For  further  information  see  the  special  chapters  on  Bacillus  trache- 
iphihis,  B.  carotovorus,  Bact.  hyacinthi,  etc.  Tests  may  also  be  made  in  air  dried  over 
sulfuric  acid  or  calcium  chloride.  Harding  &  Prucha  have  shown  recently  that 
Bacterium  campestre  remains  alive  much  longer  when  dried  on  cabbage  seed  than 
when  dried  on  glass  cover-slips.  In  their  experiments  this  organism  was  dead  on 
glass  at  the  end  of  -ten  days,  but  alive  on  seed  at  the  end  of  thirteen  months. 

EFFECT   OF   DIRECT  SUNLIGHT. 

The  exposures  should  be  made  in  a  thin  stratum  of  nutrient  agar,  not  sowed 
too  thickly  (there  may  be  several  hundred  colonies  on  the  plate,  if  properly  distrib- 
uted), in  thin-bottomed  Petri  dishes,  to  an  unclouded  sun  for  5,  10,  15,  30, 45,  and  60 
minutes,  a  portion  of  the  bottom  of  the  plate,  which  is  placed  uppermost,  being 
covered  by  some  substance  impervious  to  light,  such  as  several  folds  of  Manila  paper 


Fig.  61*  Fig.62.t 

or  of  the  black  paper  which  comes  wrapped  around  photographic  dry  plates,  covered 
in  turn  by  white  paper.  Exposures  of  several  hours  are  not  recommended.  If  the 
layer  of  agar  is  very  deep,  or  if  the  sowings  are  too  thick,  some  organisms  will  screen 
others  and  all  will  not  be  killed.  Ten  cubic  centimeters  is  a  proper  amount  of  agar  to 
use  for  a  plate  having  an  area  of  60  square  centimeters.  The  latitude,  altitude,  time  of 
year,  time  of  day,  and  intensity  of  the  light  should  also  be  recorded.  In  the  summer- 
time it  is  very  important  that  the  exposures  should  be  made  on  blocks  of  ice  or, 


*Fic.  61. — Gelatin  culture  of  Bacillus  amylovorus  (Burrill)  Trev.  in  a  Petri  dish.  Exposed  in 
1896  to  direct  sunlight  for  four  hours  on  ice  after  covering  portions  of  the  plate  with  pasteboard 
figures.  The  bacteria  grew  only  under  the  protected  parts.  Drawn  from  a  photograph  made  after 
five  days  incubation  of  the  culture  at  about  24°  C.  The  temperature  of  the  gelatin  during  exposure 
was  about  25°  C.  Three-fifths  natural  size. 

fFic.  62. — Agar  culture  of  Bacterium  phascoIiCErw.  Sm.)  in  a  Petri  dish.  Right  one-half  ex- 
posed to  direct  sunlight  for  thirty  minutes,  on  ice,  the  other  half  protected  by  several  folds  of  Manila 
paper.  Dish  then  set  away  in  the  dark  for  several  days.  One-half  natural  size.  The  scattering 
colonies  on  the  right  side  undoubtedly  grew  from  bacteria  which  were  sheltered  from  the  direct 
rays  of  the  sun  by  overlying  organisms,  »'.  e.,  the  plate  was  sown  too  thickly. 


72  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

better,  on  larger  Petri  dishes  filled  with  pounded  ice ;  otherwise,  in  case  of  30  to  60 
minute  exposures,  the  temperature  may  rise  nearly  or  quite  to  that  of  the  thermal 
death-point  of  the  organism,  and  then  we  shall  have  the  effect  of  heat  complicating 
that  of  light.  To  avoid  errors  it  is  always  best  to  take  one-half  of  each  dish  as  a 
check  (rather  than  the  whole  of  a  separate  dish),  and  the  rise  of  temperature  should 
be  carefully  recorded.  In  some  tests  made  by  the  writer  in  Washington  in  May  the 
temperature  of  the  plates  exposed  in  the  open  air  to  the  sun  for"45  minutes  (without 
ice)  rose  from  25°  to  51°  C.  Figs.  61  and  62  show  the  effect  of  sunlight  upon  thin 
sowings  of  Bacillus  amylovorus  and  Bacterium  phaseoli  in  poured-plate  (Petri-dish) 
cultures. 

VITALITY   ON   VARIOUS   MEDIA. 

By  this  I  mean  the  determination  of  the  resistance  of  organisms  to  their  own 
decomposition  products.  This  varies  greatly.  Much  may  be  learned  by  the  study 
of  old  cultures.  Do  not  discard  test-tube  cultures  until  after  many  weeks.  Examine 
frequently.  Make  transfers  from  tubes  which  have  been  inoculated  for  a  year  or 
more.  Determine  whether  this  vitality  is  due  to  spores  or  persists  in  the  ordinary 
vegetative  rods.  On  what  kinds  of  media  does  a  particular  organism  live  longest? 
Can  length  of  life  be  increased  by  occasionally  neutralizing  decomposition  products 
(acids)  with  sterile  carbonate  of  lime  ?  or  by  occasional  additions  of  food  ?  Some 
bacteria  are  veritable  revelers  in  filth  ;  others  are  extremely  sensitive  ;  all  are  soon 
under  abnormal  conditions  in  our  culture-tubes. 

Another  way  of  keeping  bacteria  alive  for  a  long  time  is  by  reducing  their  growth 
to  a  minimum.  Stock-cultures,  especially  of  perishable  organisms,  should,  generally 
speaking,  be  kept  in  the  ice-box  at  temperatures  under  15°  C.  This  greatly  reduces 
the  always  heavy  burden  of  keeping  alive  cultures  of  organisms  which  are  not 
in  immediate  demand  for  actual  experiment.  Some  will  also  remain  alive  a  long 
time  when  sealed  airtight.  Particular  organisms  may  be  kept  a  long  time  in  par- 
ticular media,  e.  g.,  Bacterium  vascularum  in  diluted  peptonized  cane-juice  gelatin, 
Bact.  Stewarti  in  milk,  etc.  Some  organisms  are  quite  resistant  to  their  own 
decomposition  products,  e.  g.,  Bacillus  coli,  Bact.  pericarditidis.  In  the  cool  box 
B.  coli  will  often  live  a  year  in  agar  stab  cultures. 

MIXED   CULTURES   AND   MIXED   INFECTIONS. 

The  behavior  of  mixed  cultures  and  mixed  infections  may  be  tested  in  various 
fluids,  making  poured  plates  from  time  to  time ;  in  tubes  of  agar,  potato,  and  other 
solid  media ;  in  crossed  streaks  on  agar  or  gelatin  plates ;  and  in  the  plants  themselves. 

When  two  bacteria,  or  a  bacterium  and  a  fungus,  are  sown  together  in  a  culture- 
medium,  there  may  be  (i)  antagonism,  with  the  crowding  out  of  one  species  ;  (2)  a 
more  or  less  complete  indifference,  both  organisms  growing  well ;  or  (3)  a  distinctly 
favorable  effect,  i.  e.,  a  marked  increase  in  growth  or  in  pathogenic  effect  due  to 
the  presence  of  the  second  organism.  The  antagonism  may  result  in  the  prompt 
destruction  of  one  of  the  organisms,  or  only  in  a  retardation  or  inhibition  which 
finally  disappears  after  the  first  organism  has  made  its  growth  and  subsided.  In 
some  cases  the  favorable  effect  of  one  organism  upon  another  is  due  to  the  fact  that 
it  prepares  food  for  it  out  of  an  unfavorable  substratum,  e.g.,  maltose  from  starch. 


BEHAVIOR   OF    MIXED    CULTURES.  73 

In  the  plant  one  organism  often  paves  the  way  for  others  which  complete  the 
destruction,  e.  g.,  Bacterium  campestre  and  Bact.  solanacearum  are  often  followed  by 
soft  white  rots.  Some  of  the  latter,  however,  are  able  to  make  their  way  unaided, 
a  fact  observed  and  known  to  the  writer  for  a  white  rot  of  the  cabbage  as  long  ago 
as  1896. 

The  simplest  way  of  studying  the  antagonistic  action  of  bacteria  is  by  means 
of  crossed  streaks  on  agar  or  gelatin  plates.  These  may  be  made  either  simulta- 
neously, or  one  after  the  other  has  begun  to  develop.  The  action  of  the  antagonistic 
organism  may  also  be  obtained  by  letting  its  products  diffuse  through  a  collodion 
sac  into  bouillon  inoculated  with  the  other  organism.  In  practice,  the  bottom  of  a 
test-tube  is  removed  and  a  collodion  sac  is  securely  fastened  in  its  place.  This  tube 
is  filled  with  the  usual  quantity  of  bouillon  and  lowered  into  a  larger  receptacle 
(tube  or  flask),  the  collodion  part  being  surrounded  by  bouillon.  The  inner  and  outer 
receptacles  are  now  plugged  with  absorbent  cotton,  and  the  apparatus  is  sterilized 
in  the  steamer  or  autoclave.  The  two  tubes  are  then  inoculated  simultaneously,  or 
the  outer  one  some  hours  or  days  after  the  inner  one.  (See  an  interesting  paper  on 
Antagonism,  by  Frost,  in  Jour.  Infect.  Diseases,  Vol.  I,  1904,  pp.  599-640).  Frost 
has  also  devised  two  new  methods  for  studying  this  subject,  viz,  the  divided-plate 
method  and  the  agar-block  method.  The  first  is  a  modification  of  the  ordinary 
streak  method.  It  is  managed  as  follows :  A  Petri  dish  is  divided  into  two  equal 
parts  by  means  of  a  glass  rod  fastened  to  the  bottom  with  collodion.  A  tube  of 
melted  agar  is  inoculated  with  the  antagonistic  organism  and  poured  into  one  half 
of  this  plate.  Into  the  other  half  sterile  agar  is  poured.  Streaks  of  the  other 
organism  are  now  made  crosswise  of  the  hardened  surface.  If  there  is  marked 
antagonism  there  will  be  a  decided  difference  in  the  behavior  on  the  two  sides  of  the 
plate,  i.  e.,  on  the  sterile  agar  as  compared  with  the  inoculated.  To  insure  a  uniform 
streak  the  inoculated  loop  should  be  swept  across  one  half  of  the  plate,  then  re- 
inoculated  and  swept  across  the  other  half  of  the  plate. 

The  method  by  agar-blocks  consists  in  substituting  agar-walls  for  collodion 
walls.  A  sterile  3-cm.-deep  Petri  dish  is  poured  full  of  nutrient  agar.  When  it  has 
solidified  it  is  cut  into  rectangular  blocks,  i  by  i  by  3  centimeters,  using  a  sterile  knife 
and  taking  all  possible  precautions  to  avoid  contamination  by  air-borne  organisms. 
A  platinum  needle  is  now  dipped  into  a  culture  of  the  supposed  antagonistic  organ- 
ism and  thrust  into  the  block  lengthwise  but  not  entirely  through  it.  The  mouth  of 
the  needle-track  is  sterilized  and  sealed  by  touching  it  for  a  moment  with  a  red-hot 
iron.  The  head  of  a  small  wire  nail  set  into  a  suitable  handle  will  answer  the 
purpose.  The  block  is  picked  up  with  sterile  forceps  and  dropped  into  a  tube  of 
sterile  bouillon,  which  then  may  be  inoculated  with  the  other  organism.  More  than 
one  block  and  tube  should  be  inoculated,  and  it  is  best  to  test  the  sterility  of  the 
outer  surface  of  the  agar-block  by  delaying  the  inoculation  of  the  bouillon  for  a  day 
or  two  after  the  inoculated  agar-block  has  been  dropped  into  place. 

Still  another  method  has  been  described  by  Frankland  and  Ward.  They  use 
the  walls  of  a  Chamberland  filter  to  keep  the  bacteria  separate.  Bouillon  for  the  one 


74  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

organism  is  placed  in  a  flask  or  large  tube.  That  for  the  other  organism  is  placed 
inside  a  Chamberland  filter,  which  is  then  sunk  into  the  other  receptacle,  whereupon 
it  is  sterilized  and  inoculated  as  in  the  collodion-sac  method. 

The  favorable  influence  of  a  second  organism  may  be  studied  in  crossed  streaks 
on  sterile  raw  potato,  carrot,  turnip,  etc.;  on  starch  jelly  ;  or  on  agar,  gelatin,  or 
silicate  jelly  with  addition  of  varying  amounts  of  the  different  plant  acids,  or  plant 
juices,  or  other  vegetable  substances.  Frost's  divided  Petri  dish  may  be  used  for 
the  jellies. 

REACTION  TO  ANTISEPTICS  AND  GERMICIDES. 

Antiseptic  has  been  defined  recently  by  Duclaux  as  follows :  Any  substance 
the  intervention  of  which  modifies  in  any  form  whatsoever  the  march  of  the  phe- 
nomena (Bibliog.,  XX,  Fermentation  alcoolique,  p.  461). 

I  still  use  the  word  with  its  old  primary  meaning  (anti,  against,  and  sepsis, 
decay).  In  this  sense  an  antiseptic  is  any  substance  which  prevents  the  multi- 
plication of  bacteria  in  putrescible  substances.  Large  doses  of  antiseptics  often 
exert  a  germicidal  action,  but  such  action  does  not  necessarily  follow.  Often  when 
the  antiseptic  substance  is  removed  or  diluted  beyond  a  certain  point  growth  takes 
place.  The  first  seven  substances  mentioned  below  possess  very  active  germicidal 
powers  and  are  antiseptic  in  correspondingly  small  doses ;  the  remainder  are  more 
or  less  valuable  antiseptics,  but  are  not  valuable  germicides. 

(i.)  Mercuric  chloride.  (5.)  Lysol.  (  9.)  Benzoic  acid. 

(2.)  Sulphate  of  copper.  (6.)  Trikresol.  (10.)   Salicylic  acid. 

(3.)  Formaldehyd  (formalin).  (7.)  Methyl  violet  (Pyoktanin).         (11.)   Chloroform. 

(4.)  Phenol  (carbolic  acid).  (8.)  Thymol.  (12.)   Sulphuric  ether. 

This  list  may  be  extended  indefinitely.  The  student  should  consult  valuable 
digests  in  Sternberg's  Text  Book  of  Bacteriology  and  in  Miquel  &  Cambier's  Traite 
de  Bacte'riologie.  Some  caution  must  be  used  in  drawing  conclusions  from  experi- 
ments. Mercuric  chloride  does  not  always  destroy  when  the  culture  medium 
contains  albuminoid  substances.  Sulphate  of  copper  is  more  active  in  water  than 
in  bouillon.*  Some  organisms  will  grow  in  a  solution  saturated  with  thymol  (e.g., 
in  bouillon).  Others  will  grow  in  the  presence  of  chloroform  (5  cc.  of  chloroform  in 
test-tubes  with  10  cc.  of  milk  or  beef-bouillon).  Ten  organisms  have  been  found  by 
the  writer  which,  under  the  conditions  named,  grew  in  the  presence  of  chloroform 
and  two  which  grew  vigorously  in  the  presence  of  thymol.  Russell  reports  one 
capable  of  growing  in  the  presence  of  sulphuric  ether.  It  is,  therefore,  not  always 
safe  to  depend  on  these  substances  as  antiseptics.  Newcombe  has  made  the  same 
observation  (Cellulose  Enzymes,  Annals  of  Botany,  Vol.  XIII,  1899,  p.  60).  In  the 
opinion  of  the  writer  the  statements  of  physiologists  respecting  the  existence  of 
enzymes  in  the  tissues  and  fluids  of  the  higher  plants  and  animals  must  be  taken 
with  much  allowance  when  chloroform,  thymol,  and  similar  antiseptics  have  been 

*Moore,  George  T.,  and  Kellerman,  Karl  F.  A  Method  of  Destroying  or  Preventing  the  Growth 
of  Algae  and  Certain  Pathogenic  Bacteria  in  Water  Supplies.  U.  S.  Department  of  Agriculture. 
Bureau  of  Plant  Industry,  Bulletin  64,  1904,  pp.  44;  see  also  Bull.  76,  Bureau  of  Plant  Industry. 
Certain  pathogenic  bacteria,  such  as  Vibrio  cholerae  and  Bacillus  typhosus,  are  destroyed  within  a 
few  hours  in  water  containing  traces  of  copper  salts  or  dissolved  particles  of  metallic  copper. 


PLATE  8. 


A  thermostat-room. 

In  the  center  of  the  bidding  and  lighted  by  electricity.     Ventilated  in  the  isrne  way  a  the  photographic  dark-roonu,  i.  e.,  by  an  exhaust-Ian 
run  by  an  electric  motor.     Three  of  the  thermortaU  were  made  by  Bauich  8t  Lorob.  the  fourth  (felted)  a  a  Rohrbeck. 


ENZYMES.  75 

depended  upon  to  keep  the  solutions  free  from  bacteria.  This  has  been  the  case 
very  frequently,  and  in  several  places  in  Greene's  interesting  book  on  Fermentations, 
published  in  1899,  it  is  said  or  inferred  that  the  addition  of  chloroform  will  prevent 
the  growth  of  bacteria.  This  might  or  might  not  be  true ;  much  would  depend 
on  the  kind  of  organisms  present.  The  medium  to  which  chloroform  or  thymol 
has  been  added  must  be  shut  in  and  shaken  continuously  if  the  full  antiseptic  value 
of  these  substances  is  to  be  obtained. 

THERMAL  RELATIONS. 

The  student  should  determine — 

(1)  Maximum  temperature  for  growth  (thermostat). 

(2)  Minimum  temperature  for  growth  (ice-box). 

(3)  Optimum  temperature  for  growth  (room  or  thermostat). 

(4)  Thermal  death-point  (ten  minutes  exposure  in  the  water-bath,  in  thin- 
walled  test-tubes  of  resistant  glass  having  a  diameter  of  16  to  17  mm.,  ordinarily  in 
10  cc.  of  moderately  alkaline  peptonized  beef-bouillon,  viz, +  15  of  Fuller's  scale). 

(5)  The  effect  of  freezing  (exposure  to  liquid  air  or  to  pounded  ice  mixed  with 
coarse  salt). 

Thermal  relations  are  among  the  most  interesting  and  should  be  studied  with 
great  care  in  case  of  every  organism.  They  offer  valuable  means  of  differentiation 
and  also  very  useful  suggestions  as  to  geographical  distribution  and  habitat.  Good 
thermostats  are  made  by  various  people.  Several  items  of  construction  are  important. 
The  water  or  oil  jacket  should  be  of  considerable  volume  (thickness)  so  as  not  to 
change  temperature  quickly ;  the  cover  should  be  thick  and  of  the  best  non- 
conducting substances.  The  opening  for  the  thenno-regulator  should  be  at  least  i  ^ 
inches  in  diameter  (so  as  to  take  a  Roux  metal-bar  thenno-regulator) ;  the  warm 
chamber  should  be  of  good  size ;  the  space  beneath  should  be  high  enough  between 
floors  to  accommodate  any  pattern  of  safety  burner;  and  last,  but  not  least,  the 
workmanship  should  be  of  the  very  best  quality,  so  that  the  apparatus  will  not 
leak.  Nearly  every  worker  has  probably  had  experience  with  leaky  thermostats  at 
some  time  in  his  life  and  knows  what  a  vexation  of  spirit  they  cause,  particularly 
if  filled  with  oil.  A  very  excellent  kind  of  thermostat  is  the  old,  large-pattern,  felt- 
covered  instrument  devised  by  Dr.  Hermann  Rohrbeck  and  figured  in  the  lower 
right-hand  corner  of  plate  8.  This  plate  shows  a  thermostat  room  with  four  thermo- 
stats in  use.  All  are  provided  with  Roux  metal-bar  thermo-regulators  and  Koch 
safety  burners.  One  is  for  quick  shifts  as  needed  ;  and  others  are  generally  kept  at 
30°,  37>40,  and  40°  or  43°  C.  These  temperatures,  in  conjunction  with  the  cool 
boxes,  thermal  baths,  and  various  room  temperatures,  enable  one  to  quickly  determine 
the  thermal  relations  of  an  organism.  The  height  of  the  room  is  10  feet,  its  depth 
7  feet,  and  its  breadth  5  feet  3  inches.  A  larger  room  would  be  more  convenient. 
Such  a  room  should  be  located  and  constructed  so  as  to  be  as  little  subject  as 
possible  to  external  changes  of  temperature.  It  should  be  lined  with  asbestos  and 
sheet  iron,  and  efficient  safety  burners  should  be  used  to  the  exclusion  of  all  others 
(see  L/autenschlager's  catalogue).  The  improved  Koch  safety  burner  is  probably  the 
best  All  burners  require  frequent  inspection. 


76 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


The  writer  has  no  very  satisfactory  way  of  making  exposures  for  determining 
the  minimum  temperature  for  growth.  His  method  is  to  make  such  exposures  in 
the  bottom  of  a  large,  well-filled  ice-box,  which  is  opened  as  little  as  possible  during 
the  progress  of  the  tests,  and  then  only  for  the  briefest  periods.  The  degree  of  cold 


Fig.  63* 

is  governed  by  the  amount  of  ice.  A  good  thermometer  is  exposed  in  the  midst  of 
a  bundle  of  inoculated  tubes,  and  if  the  temperature  shows  any  tendency  to  rise 
more  ice  is  added.  Under  the  most  favorable  circumstances  the  temperature  of  the 

*Fic.  63. — Modification  of  the  Ostwald  water -bath  used  by  the  writer  for  thermal  death-point 
experiments.  This' consists  of  a  porcelain-lined  pot  n  inches  in  diameter  at  the  top.  This  is  filled 
with  water  kept  in  motion  by  a  water-wheel  turned  by  electricity.  The  heat  is  applied  by  means  of  a 
Friedburg  burner  and  is  controlled  by  Roux's  thermo-regulator.  Murrill's  gas-pressure  regulator 
is  shown  at  the  left. 


THERMAL    RELATIONS. 


77 


air  in  the  bottom  of  the  chest  may  be  kept  fairly  constant  for  some  days  or  weeks,  but 
with  marked  external  fluctuations  of  temperature  trustworthy  results  can  be  obtained 
only  by  constantly  watching  the  box. 
What  one  needs  for  this  work  is  a  good- 
sized  room  kept  at  o°  C.,  or  a  little  below, 
in  which  thermostats  may  be  installed  at  GV 
temperatures  a  little  above  freezing,  e.  g., 
+  2°,  +5°,  +7°,  etc.  It  would  then  be 
very  easy  to  determine  the  minimum 
temperature  at  which  any  organism  will 
grow — as  easy  as  it  is  now  to  determine  the 

maximum.     Different  levels  in  the  same  room  may  afford  constant 
and  useful  differences  in  temperature. 

The  thermal  death-point,  which  is  a  purely  arbitrary  standard, 
depending  on  the  age  and  kind  of  culture,  its  volume,  and  the  length 
of  exposure,  as  well  as  the  temperature,  is  when  properly  determined 
not  least  valuable.  The  writer,  following  that  one  of  Dr.  Sternberg's 
methods  which  is  easiest  to  carry  out,  uses  10  cc.  portions  of  moder- 
ately alkaline  (-f-io  or  +15)  peptonized  beef-brothf  in  test-tubes  of 
uniform  diameter  (16  to  17  mm.),  inoculates  from  recent  bouillon- 
cultures  with  care  not  to  touch  the  sides  of  the  tube  above  the  fluid, 
thrusts  the  tubes  deep  into  the  hot  water,  and  exposes  for  ten  minutes. 
All  who  make  this  test  are  urged  to  use  standard  alkaline  beef- 
bouilloii  (for  all  organisms  growing  well  in  this  medium)  and  to 
limit  the  exposure  to  exactly  ten  minutes,  so  that  easy  comparisons 
may  be  made.  The  five  minutes  exposure  which  has  been  recom- 
mended by  some  authors  is  rather  too  short,  since  it  only  a  little 
more  than  suffices  to  warm  the  fluid  up  to  the  required  temperature. 
Inoculation  while  the  tubes  are  in  the  bath  and  after  the  fluid  has 
been  brought  to  the  required  temperature  is  inconvenient  and  has  no 
special  advantage. 


Fig.  64.* 


*Fic.  64.— Roux's  thermo-regulator,  made  by  Maison  Wiesnegg  (P.  Lequeux),  Paris.  The  parts 
requiring  description  are  as  follows:  A,  bar  composed  of  two  metals  (which  expand  and  contract  un- 
equally) attached  at  bottom  and  free  at  the  top,  which  moves  with  increased  heat  in  the  direction  of 
the  arrow;  B,  arm  on  which  the  upper  part  of  the  apparatus  moves  freely  when  K  is  turned;  C, 
stiff  spring;  D,  long  rod  which  controls  the  gas-inflow,  and  the  spring  movement  of  which  is  in 
the  direction  of  the  arrow  except  when  controlled  by  the  counter  movement  of  A,  due  to  lessened 
heat;  E,  gas-inflow;  F,  gas-chamber,  of  glass;  G,  gas-outflow,  to  the  burner;  H,  rubber  stopper; 
I,  cylinder  screwing  into  L,  and  provided  with  capped  upright  tube  filled  with  vaseline  to  prevent 
gas  from  escaping  in  the  direction  of  D.  The  button  shown  in  the  gas-chamber  at  the  left  is  part 
of  D,  and  the  gas  enters  the  chamber  between  it  and  the  left  end  of  L,  the  size  of  the  opening,  and 
consequently  the  amount  of  gas,  varying  with  the  slightest  movement  of  A.  Different  temperatures 
are  obtained  by  turning  the  button  K.  The  constant  gas-flow  is  provided  for  by  a  small  opening  on 
the  lower  side  of  L  at  its  extreme  left,  in  the  gas-chamber.  About  two-fifths  actual  size. 

fThe  thermal  death-point  in  acid  media  is  considerably  higher — at  least  that  of  several  organ- 
isms which  have  been  tested  in  the  author's  laboratory. 


78  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

An  excellent  water-bath  is  that  known  as  the  Ostwald-Pfeffer.  The  experi- 
menter may,  however,  construct  one  for  himself  out  of  a  medium-sized,  thick- 
walled,  porcelain-lined  iron  kettle  (fig.  63).  This  should  rest  on  a  ring  of  heavy 
strap-iron  supported  by  four  stout  iron  legs.  The  burner  required  may  be  Dr. 
Friedburg's  safety  burner  (a  very  inexpensive  and  good  pattern).  The  thermo- 
regulator  may  be  a  common  Reichert  if  the  mercury  seal  is  cleaned  from  oxide 
frequently.  In  such  regulators  a  sharper  contact  and  a  longer  freedom  from  obstruc- 
tion is  said  to  be  obtained  (Dr.  Harris)  by  putting  a  drop  of  olive  oil  on  top  of 
the  mercury.  A  much  better  instrument  is  the  metal-bar  mechanism  known  as  the 
Roux  regulator  (fig.  64).  This  may  be  procured  from  the  Maison  Wiesnegg,  in 
Paris.  It  should  be  kept  from  direct  contact  with  the  water  and  consequent  rusting 
by  burying  it  in  a  close-fitting  glass  tube  filled  with  olive  oil  or  glycerin.  This 
tube  is  then  sunk  deep  into  the  water  and  clamped  to  the  wall  of  the  kettle,  which 
should  have  perpendicular  sides.  The  water  is  kept  in  motion  by  means  of  a  hori- 
zontal paddle-wheel  at  the  bottom  of  the  kettle.  This  consists  of  four  light,  oblique 
zinc  or  copper  vanes  (nickeled  copper  is  preferable)  soldered  to  a  long  central  rod 
which  fits  into  a  socket,  below,  and  near  its  upper  end  passes  through  a  hole  or  loop 
in  a  horizontal  metal  arm  (a  foot  or  less  above  the  kettle),  the  other  end  of  which  is 
clamped  to  the  upright  rod  of  a  solid  iron  tripod,  or  fastened  to  a  rod  bolted  to  the 
table.  If  compressed  air  can  be  had,  a  stiff  cardboard  windmill  fastened  to  the  upper 
end  of  the  vertical  rod  completes  the  mechanism.  The  central  part  of  the  wind- 
wheel  may  be  of  cork.  The  vertical  rod  may  be  a  piece  of  glass  tubing,  in  which 
case  it  is  cemented  into  a  socket  of  the  short  metal  post  to  which  the  vanes  of  the 
water-wheel  are  soldered.  If  a  wind-wheel  is  attached,  it  is  more  convenient  to  have 
the  vertical  rod  in  two  parts,  fastened  by  a  coupling.  The  rod,  with  its  water-wheel 
attachment,  may  also  be  turned  by  some  electrical  device.  The  latter  is  the  most 
convenient  method.  In  fig.  63  the  electric  motor  is  not  shown.  This  stands  in 
a  small  box  screwed  to  the  under  side  of  the  table  at  the  right.  The  switch  is  fastened 
to  the  wall  above  and  back  of  the  top  of  the  thermo-regulator.  The  pulley  band 
is  of  smooth  rounded  leather  one-eighth  inch  in  diameter.  The  electric  current  is 
passed  through  an  Edison  lamp  screwed  under  the  table  to  reduce  the  velocity  of 
the  motion.  With  the  lamp  in  place  and  the  current  cut  down  to  the  minimum 
the  number  of  revolutions  per  minute  is  55,  and  the  temperature  of  the  water  is  the 
same  in  all  parts  of  the  bath.  The  simplest  contrivance  of  all  is  to  make  the 
water-wheel  and  upright  shaft  of  wood,  to  be  turned  by  hand. 

In  localities  where  the  gas-pressure  is  exceedingly  variable,  Paul  Murrill's  gas- 
pressure  regulator  (at  the  left  in  fig.  63)  will  be  found  useful.  This  is  made  by 
Eberbach  &  Co.,  Ann  Arbor,  Mich,  (see  Journal  of  Applied  Microscopy, Vol.  I,  p.  92, 
or  Centralb.  f.  Bakt,  i  Abt,  Band  XXIII,  1898,  p.  1056.)  The  gas-pressure  may  be 
somewhat  improved  by  simply  passing  the  gas  through  a  big  bottle  (see  top  of 
thermostat  311  in  plate  8).  The  Anschiitz  normal  thermometers,  with  long  stem 
and  scale  divided  into  fifths,  are  very  convenient  for  determining  temperatures 
(fig.  65).  They  come  in  sets  of  seven,  but  may  also  be  had  separately.  The  most 
frequently  useful  are  No.  i  (scale  — 15°  to  +55°)  and  No.  2  (scale  +  45°  to  +  105). 


THERMAL   RELATIONS.  79 

They  cost  9  marks  each  when  ordered  direct  from  Berlin,  and  can  be  had  without 
delay.     Good  American  thermometers  are  made  by  Henry  Green,  New  York. 

With  this  open  bath  it  is  easy  to  keep  the  range  of  temperature  down  to 
o.i  to  0.3  of  a  degree,  and  the  writer  has  frequently  exposed  tubes  for  ten  minutes 
without  appreciable  change  in  temperature.  Temperatures  may  be  read  easily  to 
o.  i  degree  by  means  of  a  Zeiss  aplanat  lens  magnifying  six  times  (fig.  25), 
and  should  be  recorded  for  each  half  minute  during  the  exposure.  Under 
no  circumstances  should  exposures  be  made  in  water  which  is  not  agitated. 
Of  course,  for  accurate  reading  the  eye  and  the  center  of  the  lens  must  be 
level  with  the  top  of  the  column  of  mercury.  The  lens  may  be  supported 
at  the  proper  level  on  a  grooved  piece  of  cork.  If  possible  the  thermom- 
eter used  should  be  compared  with  some  standard  instrument.  If  not,  it 
should  at  least  be  compared  with  several  other  good  thermometers  in  the 
same  laboratory.  The  test-tubes  are  supported  by  perforated  corks  thrust 
into  holes  bored  through  a  rectangular  piece  of  hard,  heavy  wood. 

The  writer  formerly  made  use  only  of  the  first  four  tests.  It  seemed 
hardly  worth  while  to  recommend  that  all  bacteria  be  tested  for  the  killing 
effect  of  cold,  so  long  as  we  had  nothing  but  the  inconvenient  and  more 
or  less  inexact  methods  of  salt  and  pounded  ice  or  of  ether  and  frozen 
CO* ;  but  now  that  liquid  air  may  be  obtained  at  a  small  price  in  many 
of  the  larger  cities,  can  be  shipped  long  distances,  and  can  be  used  with 
so  little  inconvenience,  there  is  no  good  reason  why  the  effect  of  freezing 
should  not  be  determined  in  all  cases,  since  in  some  instances  it  is  likely 
to  prove  a  valuable  means  of  differentiation.  The  bacteria  may  be  exposed 
in  5  cc.  portions  of  distilled  water  or  bouillon  in  block-tin  test-tubes,  or 
preferably  in  tubes  of  resistant  glass,  for  standard  periods,  e.  g.,  one-half 


hour,  i  hour,  6  hours,  12  hours,  24  hours,  48  hours,  etc.     They  may  also 


be  exposed  to  alternate  freezing  and  thawing  every  fifteen  minutes  or 
thirty  minutes  until  all  are  dead.  To  avoid  endospores,  the  depressing 
effect  of  by-products,  etc.,  young  cultures  should  be  used,  and,  of  course, 
all  should  be  of  the  same  age  and  grown  in  the  same  medium,  i.  e., 
bouillon  cultures  24  hours  or  48  hours  old.  The  tests  should  be  quanti- 
tative rather  than  qualitative.  They  may  be  made  as  follows  :  Into  5  cc. 
of  sterile  water  or  standard  bouillon  a  carefully-measured  quantity,  i.  e., 
one  loop,  5  drops,  ^  cc.,  etc.,  of  the  culture  is  placed,  stirred  very  thor- 
oughly, and  allowed  some  time  for  diffusion.  To  avoid  zooglcese,  which 
form  early  in  some  species,  and  to  reach  more  uniform  measurements, 
it  is  recommended  to  take  the  loop  from  a  bouillon  culture  rather  than 
from  agar  or  other  solid  media.  After  sufficient  time  has  elapsed  for 
Fig.  65*  unifonn  diffusion,  six  Petri-dish  poured  plates  are  made  from  each  of  the 

inoculated  tubes.     The  plates  should  be  of  the  same  diameter  (area  of  60  sq.  cm.). 

The  amount  of  agar  used  for  each  plate  should  be  10  cc.,  and  the  amount  of  infec- 

*Fic.  65. — Anschiitz  normal  thermometer  with  degrees  divided  into  fifths   (Centigrade  scale). 
For  use  in  thermal  death-point  tests.    About  three-fourths  actual  size. 


8o 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


tious  material  used  should  be  the  thinnest  obtainable  film  of  fluid  across  a  carefully- 
measured  i  mm.  loop,  so  as  to  avoid  crowding  the  plates.  The  same  loop  should 
be  used  in  all  cases,  and  it  should  be  dipped  into  and  out  of  the  fluid  always  in 
the  same  way.  After  pouring,  set  the  plates  on  a  perfectly  level  spot  (fig.  66),  until 
the  agar  has  hardened.  If  the  work  has  been  well  done,  there  should  develop 
an  approximately  uniform  number  of  colonies  in  each  plate.  The  tubes  of  inocu- 
lated water  or  bouillon  are  then  immediately  lowered  into  the  liquid  air  and  exposed 
to  it  for  the  predetermined  time,  after  which  six  additional  Petri-dish  poured  plates, 
of  the  same  size  and  inoculated  in  the  same  way,  are  made  from  each  tube  for 
comparison  with  those  prepared  before  the  exposure.  The  tubes  may  be  thawed  out 
by  exposure  to  the  air  for  three  minutes  and  then  to  tap-water  for  five  to  seven 
minutes.  The  exposures  are  best  made  in  Dewar  glasses  (fig.  67).  When  the 
exposures  are  long,  a  loose  tuft  of  absorbent  cotton  should  be  placed  in  the  mouth 
of  the  glass,  or  it  should  be  covered  with  a  hair-cloth  cap,  to  prevent  excessive 


Fig.  66* 

evaporation.  Under  these  conditions  the  air  remains  liquid  for  a  number  of  days. 
At  first  the  temperature  is  about  minus  190°  C.,  rising  gradually  to  minus  180°  C., 
since  the  nitrogen  evaporates  somewhat  faster  than  the  oxygen.  The  glasses  are 
fragile  and  should  be  handled  carefully,  especially  when  filled  with  the  air.  As  long 
as  they  contain  liquid  air  it  is  safer  to  keep  them  in  their  containing-case,  packed 
about  with  cotton  or  felt.  One  should  be  careful  to  avoid  cracking  the  inner  wall 
of  the  glass,  as  might  happen  by  dropping  some  hard  substance  into  the  receptacle, 
otherwise  an  explosion  will  occur,  the  space  between  the  two  walls  of  the  Dewar 
glass  being  a  very  perfect  vacuum. 

When  the  exposures  are  made  in  block-tin  tubes,  the  culture  should  be  frozen 
at  once  on  pouring  into  the  tube  and  the  second  set  of  plates  should  be  made 
as  soon  as  the  fluid  has  thawed,  /'.  ^.,  within  about  ten  minutes,  for  which  purpose 
the  culture  should  be  poured  out  into  a  glass  tube,  otherwise  complications  due  to 

*Fic.  66. — Leveling  (nivelling)   apparatus  for  use  in  making  poured  plates.     About  one-third 
actual  size. 


THERMAL    RELATIONS. 


81 


the  gennicidal  action  of  the  metal  might  arise.  In  no  case  should  the  cultures  be 
incubated  in  the  tin  tubes.  When  exposures  are  made  in  test-tubes  of  resistant 
Jena  glass,  the  cultures  must  be  lowered  into  the  liquid  air  gradually,  the  fluid  being 
frozen  from  the  bottom  upward  to  avoid  cracking  the  tubes.  It  requires  about  four 
minutes  to  properly  freeze  a  culture  in  a  glass  test-tube.  Large  volumes  of  culture 
media  should  not  be  lowered  into  the  liquid  air,  as  it  is  wasteful,  the  air  boiling  away 
rapidly.  The  writer  began  his  experiments  with  block -tin  tubes,  as  shown  in  fig. 
67,  but  now  uses  tubes  of  Jena  glass.  The  latter  crack  occasionally  in  spite  of  care. 


Fig.  67* 

For  very  rapid  freezing  the  amount  of  fluid  in  the  tube  may  be  reduced  to  i  cc. 
Liquid  air  in  Dewar  glasses,  and  compressed  oxygen,  hydrogen,  and  carbon  dioxide  (?) 
in  steel  cylinders  may  be  had  from  the  Eagle  Oxygen  Company,  Incorporated,  121 
West  Eighty-ninth  Street,  New  York  City.  The  tanks  of  compressed  gases  may  be 
bought  or  rented.  The  following  sizes  may  be  had  :  Fifty  gallons  (280  pounds 
pressure  per  square  inch) ;  100  gallons  (240  pounds  pressure)  ;  150  gallons  (225 
pounds  pressure)  ;  and  200  gallons  (280  pounds  pressure).  Cylinders  may  also  be 
had  with  the  gas  under  much  greater  pressure.  The  cost  of  the  oxygen  is  2}4  cents 

*Fic.  67. — Dewar  glass  for  liquid  air,  and  block-tin  test-tubes  used  in  first  low  temperature  ex- 
periments with  bacteria.     About  one-sixth  actual  size. 


82  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

per  gallon.  The  wrought-steel  cylinders  cost  about  $10  each.  A  good  quality 
of  resistant-glass  test-tubes  may  be  had  from  Greiner  &  Friedrichs,  Stiitzerbach, 
Germany.  One  sort  has  a  faint-blue  longitudinal  stripe  blown  into  the  glass, 
another  kind  has  the  letter  "  R  "  etched  on  the  upper  part  of  each  tube.  Tubes 
without  any  distinguishing  mark  should  not  be  purchased,  as  they  are  likely  to 
become  mixed  with  ordinary  non-resistant  tubes.  The  cost  of  these  tubes,  duty 
free,  is  about  $16  per  thousand.  Good  Petri  dishes  may  be  obtained  from  the  same 
firm,  and  also  from  E.  H.  Sargent  &  Co.,  Chicago. 

The  temperature  demands  of  bacteria  are  extremely  variable.     Whole  groups 
of  them  are  able  to  live  under  conditions  which  would  be  impossible  for  the  higher 


Fig.  68* 

plants  and  animals.  Many  of  the  northern  forms,  especially  those  which  grow  in 
water,  are  adapted  to  low  temperatures.  The  organisms  of  dung-heaps  and  thermal 
springs,  and  the  tropical  forms,  often  grow  at  high  temperatures. 

For  a  very  few  species  it  has  been  known  that  prolonged  freezing  or  repeated 
freezing  and  thawing  destroys  the  weaker  individuals  and  finally  all.  (See  Bibliog., 
XXXIII,  especially  papers  by  Sedgwick  &  Winslow,  and  by  Park ;  consult  also  an 
earlier  paper  by  Prudden,  Bibliog.,  XL, VI.)  For  the  bacteria  as  a  whole,  however, 
it  has  been  assumed  that  ordinary  freezing  or  even  very  intense  cold  simply  inhibits 

*Fic.  68. — Petri-dish  poured  plate  of  Bacillus  tracheifhilus.  The  10  cc.  of  nutrient  agar  was 
inoculated  with  a  carefully  measured  loop  of  a  fluid  culture.  The  fluid  culture  was  then  exposed 
in  block-tin  test-tubes  to  the  temperature  of  liquid  air,  after  which  another  plate  (fig.  69)  was  made. 


THERMAL   RELATIONS.  83 

growth  for  the  time  being.  Such  statements  have  been  based  on  certain  qualitative 
tests  and  do  not  tell  the  whole  truth.  In  the  writer's  experiments  with  liquid  air 
great  differences  have  been  detected,  the  reduction  by  exposure  for  one-half  hour 
varying  from  15  per  cent,  or  less,  to  90  per  cent,  or  more,  according  to  the  species 
tested.  Fully  50  per  cent  of  many  sorts,  grown  in  bouillon,  are  destroyed  by  a  single 
short  exposure  (see  figs.  68  and  69).  Query  :  Is  intense  cold  any  more  harmful  to 
bacteria  than  simple  freezing?  Are  young  or  old  cultures  most  susceptible? 
Are  they  killed  by  the  rupture  of  the  cell-wall  due  to  the  formation  of  ice- crystals,  or 
simply  by  the  abstraction  of  water?  Why  do  some  resist  several  freezings?  Can 
endospores  be  killed  in  this  way  ?  Consult  '01,  d'Arsonval  (Bibliog.,  XXXIII)  and 


Fig.  69* 

Smith  &  Swingle,  the  Effect  of  Freezing  on  Bacteria,  Proc.  Sixth  Ann.  Meeting 
Soc.  Am.  Bacteriologists,  December  27,  1904;  Science,  N.  S.,  Vol.  XXI,  1905,  pp. 
481-483.  For  opposing  views  see  '02,  Macfadyen,  Bibliog.,  XXXIII. 

Live  steam  acts  upon  the  growing  bacteria  very  quickly.  All  bacteria  not  in 
spore  form,  or  in  some  other  way  protected  from  the  direct  action  of  the  heat  by 
what  surrounds  them,  are  promptly  destroyed  by  steam  heat  at  100°  C.,  an  exposure 
of  a  minute  or  two  being  ample,  except,  possibly,  in  case  of  some  of  the  thermo- 


*FiG.  69. — Same  as  fig.  68,  but  made  after  exposure  for  twenty  hours  to  liquid  air.  Number 
of  colonies  reduced  two-thirds.  Exposed  in  test-tubes  of  Jena-glass  for  one-half  hour,  the  reduc- 
tion was  nearly  as  great,  i.  e.,  over  50  per  cent.  In  this  latter  case  the  agar  plates  were  incubated 
7  days  at  30°  C,  before  the  count  was  made. 


84 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


philic  species.  Usually  even  the  most  resistant  spores,  if  freely  exposed,  are  destroyed 
by  one  to  two  hours  exposure  to  150°  C.,  of  dry  heat,  or  by  thirty  minutes  exposure 
on  each  of  three  consecutive  days  to  streaming  steam  at  100°  C.  Some  very 
resistant  spores  have  survived  a  single  steaming  or  boiling  of  five  or  six  hours 
duration  (eight  hours  in  one  of  Tyndall's  experiments),  and  it  is  not  unlikely  that 
some  slowly  germinating  sorts  may  be  able  to  resist  discontinuous  steamings  for 
three  days.  It  is  possible  also  that  there  may  be  some  sorts  able  to  germinate  and 
again  assume  a  resistant  spore  form  in  less  than  twenty-four  hours  although  this  is 
not  probable.  Some  spores  are  destroyed  by  a  short  boiling  at  100°  C.,  and  all 
spores  are  quickly  destroyed  by  steam  under  pressure,  i.  e.,  in  an  autoclave.  A 


Fig.  70  * 

temperature  of  1 10°  C.  for  ten  or  fifteen  minutes  is  sufficient.  Exposure  of  media 
to  higher  temperatures  and  for  longer  periods  should  be  carefully  avoided.  It  must 
be  remembered,  however,  in  using  autoclaves,  that  all  of  the  air  must  be  replaced 
by  steam  before  the  apparatus  is  closed,  otherwise  the  temperature  to  which  the 
medium  is  exposed  will  not  correspond  to  that  indicated  by  the  pressure  gage. 
The  -most  convenient  autoclaves  known  to  the  writer  are  the  large  sizes  of  the 

*Fic.  70. — Earliest  stage  of  fruit  spot  on  green  plums,  due  to  Bacterium  pruni  (Erw.  Sm.).  The 
bacteria  have  entered  through  the  stoma.  They  disappear  farther  in,  and  also  a  few  micra  to  either 
side  of  this  stoma,  as  shown  by  an  examination  of  the  serial  sections.  Material  fixed  in  strong  alco- 
hol, infiltrated  with  paraffin,  and  cut  on  the  microtome  in  series.  Section  stained  with  carbol-fuchsin 
and  drawn  directly  from  the  microscope  with  the  aid  of  a  camera  lucida. 


PLATE  9. 


Chamberland  autoclave. 

Heat  is  applied  to  the  bottom  by  means  of  a  double  ring  of  Bunaen  burners.      No  wrench  is  required  for  fastening  on  the  top.     About 

one-eighth  natural  size. 


THERMAL   RELATIONS.  85 

pattern  designed  by  Chamberland  and  made  by  the  Maison  Wiesnegg  (P.  Lequeux), 
Paris,  France,  the  steam  being  generated  by  gas  (plate  9).  The  steam  gage  is  at 
the  left ;  in  the  middle  is  the  valve  through  which  the  hot  air  is  allowed  to  escape 
when  the  instrument  is  wanned  up  ;  at  the  right  is  the  steam  safety-valve.  The 
temperature  is  manipulated  by  regulating  this  valve.  By  leaving  the  vent  open  the 
apparatus  may  be  used  as  an  ordinary  steam  sterilizer.  It  may  also  be  used  as  a 
distilled-water  apparatus  by  attaching  a  condenser  to  the  exit  pipe  of  the  middle 
vent,  but  such  water  must  not  be  used  for  culture  media.  A  very  good  autoclave 
is  also  made  by  the  Kny-Scheerer  Co.,  New  York.  Harding  recommends  for  auto- 
claves the  use  of  steam  from  the  engine-room  boiler.  This  is  convenient,  provided 
one  can  always  have  steam  ready  during  the  summer  months.  An  autoclave,  like  a 
steam  boiler,  which  it  is,  must  be  watched  carefully  if  it  is  not  some  time  to  explode 
from  excess  of  heat  or  lack  of  water.  Each  time  before  use  one  should  see  that  the 
apparatus  contains  sufficient  water. 

Soils  are  rather  difficult  to  sterilize.  They  may  be  spread  in  thin  layers  and 
dry-heated  for  several  hours  at  150°  C.,  or  may  be  heated  in  the  autoclave  for  an 
hour  under  a  pressure  of  two  atmospheres,  taking  care  to  drive  all  the  air  out  of  the 
soil  before  closing  the  apparatus.  It  is  not  likely,  however,  that  soils  can  be  treated 
in  this  way  without  undergoing  certain  physical  and  chemical  changes.  Small 
pots  of  soil  may  be  heated  in  the  steamer  at  100°  C.  for  two  hours  on  each  of  five 
successive  days. 

The  reason  for  preparing  all  media  in  the  autoclave,  or  by  heating  in  the 
steamer  at  100°  C.  on  three  successive  days  (the  ordinary  way),  is  because  we  are 
never  certain  in  what  particular  case  resistant  spores  may  be  present.  One  short 
steaming  is  often  sufficient  to  sterilize  media  prepared  in  a  cleanly  way,  as  every 
bacteriologist  knows  who  has  had  much  experience,  but  'now  and  then,  in  spite  of 
all  care,  resistant  spores  will  find  their  way  into  culture  media,  and  for  this  reason 
it  is  best  in  all  cases  (especially  in  teaching  students)  to  adhere  to  a  routine  of  three 
steamings.  Large  masses  of  fluid  (beakers,  flasks)  require  longer  steamings  than 
test-tube  cultures.  The  writer  gives  double  time,  or  triple  time.  Discontinuous 
boiling  as  a  means  of  sterilization  was  introduced  in  1877  by  Tyndall,  who  well 
says  respecting  the  sterilization  of  liquids  :  "Five  minutes  of  discontinuous  heating 
can  accomplish  more  than  five  hours  continuous  heating."* 

Most  plant-pathogenic  bacteria  of  temperate  and  cold  regions  have  a  lower 
optimum  and  maximum  temperature  for  growth  and  a  lower  thermal  death-point 
than  species  pathogenic  to  warm-blooded  animals.  The  maximum  temperature  for 
growth  is  usually  at  or  below  36°  C.  We  should  not,  however,  expect  this  to  be 
true  of  bacterial  plant  parasites  in  tropical  and  sub-tropical  regions,  about  which, 
however,  little  is  known  beyond  the  mere  fact  that  such  parasites  occur.  Savastano 
states  that  the  optimum  temperature  for  the  olive-knot  organism,  which  is  said  to 
be  more  prevalent  at  the  southern  than  at  the  northern  limit  of  olive-growing, 

*This  method  appears  to  have  been  known  to  housewives  for  a  much  longer  time.  In  Dr.  Sam- 
uel Johnson's  Dictionary  (first  Am.  from  eleventh  London  ed.)  I  find  the  following  definition: 
"  Biscuit,  A  kind  of  hard,  dry  bread  made  to  be  carried  to  sea.  It  is  baked  for  long  voyages  four 
times." 


86 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


i.  e.,  commonest  in  southern  Italy,  Sicily,  and  Algeria,  lies  between  32°  and  38°  C. 
In  my  own  experiments  with  this  organism,  obtained  from  olive  trees  in  California, 
I  have  found  its  maximum  temperature  to  be  above  35°  and  below  37.5°  C.  The 
optimum  temperature  of  Bacterium  solanacearum,  which  is  very  destructive  to 
potatoes  and  tomatoes  in  the  southern  United  States,  is  probably  about  35°  C. — at 
least  it  grew  readily  and  remained  alive  for  a  long  time  in  bouillon  kept  at  37°  C. 
Its  maximum  temperature  is  39  +°  C.  Bacillus  carotovorus,  one  of  the  best  known 
of  the  soft-rot  organisms,  grows  well  in  the  thermostat  at  33°  to  34°  C.  Its  maxi- 
mum temperature  is  at  39°  C.  or  slightly  below  (Jones).  Bacillus  aroidete,  whose 
temperature  relations  were  recently  studied  carefully  by  Townsend,  has  a  maximum 


Fig.  71  * 

temperature  of  41°  C.  A  temperature  of  40°  C.  retards  growth,  but  does  not  prevent 
it.  This  organism  was  isolated  from  calla-lily  conns,  but  is  capable  of  causing  a 
soft  rot  in  potatoes,  carrots,  turnips,  and  many  other  plants  (fig.  102).  The  maxi- 
mum temperature  of  Bacillus  oleraceae,  recently  described  by  Harrison,  is  said  to 
be  about  42°  C.  This  causes  a  soft  rot  of  cauliflower. 

The  range  of  temperature  suited  for  the  growth  of  particular  bacteria  varies 
greatly.  Some  species  are  able  to  grow  through  a  range  of  50°  C.  Many  tolerate 
a  range  of  only  about  30°  C.  Certain  animal-pathogenic  forms  have  through  long 
subjection  to  a  peculiar  environment  become  restricted  to  a  still  narrower  range. 

*Fic.  71. — Bacterium  pruni.  Early  stage  of  a  leaf-spot  in  the  plum.  The  small  spot  was  water- 
soaked  in  appearance,  but  it  had  not  yet  collapsed.  The  bacteria,  which  are  most  abundant  in  the 
mesophyll,  undoubtedly  entered  the  leaf  through  the  stomata,  three  of  which  are  shown  in  the  section. 
Material  treated  as  in  fig.  70.  Section  drawn  with  the  aid  of  an  Abbe  camera.  It  represents  as 
nearly  as  possible  one  plane. 


THERMAL    RELATIONS.  87 

Some  bacteria  grow  well  only  in  the  cool  box,  others  only  in  the  thermostat 
at  blood-heat  or  at  higher  temperatures, — temperatures  elevated  enough  to  quickly 
destroy  the  unprotected  protoplasm  of  the  higher  plants  and  animals.  Few  of  the 
bacteria  commonly  studied  will  grow  at  temperatures  much  above  40°  C.,  but  this 
by  no  means  expresses  the  whole  truth. 

The  lowest  temperature  at  which  growth  will  take  place  ranges  in  different 
species  all  the  way  from  o°  C.,  and  probably  a  few  degrees  below  (certain  salt-water 
bacteria)  to  -\-  40°,  +  50°,  +  56°,  and  even  +  60°  C.  (certain  thermophilic  species 
found  in  dung-heaps,  hay-mows,  silos,  hot  springs,  etc.).  The  highest  temperature 
at  which  growth  will  take  place  ranges  from  as  low  as  30°  C.  (and  probably  lower*) 
to  as  high  as  75°,  or  80°  C.,  or  even  89°  C.,  according  to  Setchell.  Higher  temper- 
atures have  been  recorded,  but  I  have  here  used  only  those  determined  with  care  in 
the  exact  places  frequented  by  the  bacteria.  This  will  be  better  appreciated  if  it  is 
remembered  that  a  temperature  of  60°  C.  (140°  F.)  can  be  endured  by  the  fingers  only 
a  few  seconds,  while  70°  C.  (the  optimum  for  some  of  these  species)  is  intolerable 
to  the  hand  even  for  the  shortest  period.  It  seems  incredible,  on  first  thought,  it  is 
so  opposed  to  our  customary  observations,  that  any  organism  whatsoever  should 
be  able  to  live  at  a  temperature  only  1 1  degrees  below  the  boiling  point  of  water 
Nevertheless,  protoplasm  is  an  extremely  adaptable  substance,  and  it  is  conceivable 
that  some  organisms  might  grow  at  a  temperature  considerably  higher. 

The  thermal  death-point  (10  minutes  exposure)  ranges  from  43°  C.  for  Bacillus 
tracheiphilus,  the  lowest  yet  recorded, f  to  temperatures  only  a  few  degrees  under 
the  boiling  point  (100°  C.).  For  many  species  the  thermal  death-point  lies  between 
50°  and  60°  C.  Russell  &  Hastings  have  recently  discovered  in  milk  a  Micrococcus 
whose  thermal  death-point  is  76°  C. 

As  the  upper  and  lower  thermal  boundaries  of  growth  are  approached  some 
functions  are  extinguished  in  advance  of  others;  e.  g.,  pigment  production,  patho- 
genicity,  and  sporulation  disappear  considerably  in  advance  of  loss  of  power  to 
reproduce  by  fission. 

OTHER   HOST   PLANTS. 

Plants  of  related  species,  genera,  and  families  should  be  tested.  If  the  disease 
appears  to  be  new  to  literature,  it  is  also  especially  important  to  inoculate  those 
plants  which  have  been  reported  to  be  subject  to  bacterial  disease  and  the  nature  of 
which  disease  is  still  in  doubt.  Many  facts  of  scientific  and  economic  interest  will 
be  brought  to  light  in  this  way,  and  now  and  then  the  experimenter  may  be  able  to 
clear  away  some  of  the  fog  which,  owing  to  the  uncertain  and  contradictory  state- 
ments of  a  majority  of  our  plant  pathologists,  still  hangs  over  the  origin  and  nature 
of  most  of  these  diseases. 

Some  plant  pathogenes  appear  to  be  quite  narrowly  restricted.  They  attack 
only  one  host  plant,  or  at  most  a  few  hosts  belonging  to  related  species  or  genera. 
Others,  particularly  some  of  the  soft-rot  bacteria,  attack  many  kinds  of  plants  belong- 
ing to  widely  different  families.  The  history  of  pear-blight,  however,  shows  us  that 

*Since  this  was  written  Molisch  states  (1.  c.,  p.  93)  that  gelatin  cultures  of  his  Bacterium  phos- 
phorcum  were  dead  at  the  end  of  48  hours  when  exposed  to  a  temperature  of  30°  C.  The  maximum 
temperature  of  this  organism  is  said  to  be  about  28°  C. 

tVery  recently  Marsh  has  found  a  fish  parasite  which  is  said  to  have  a  thermal  death-point  of 
42°  C.  (See  VI,  Bibliography  of  General  Literature.) 


88 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


the  restriction  of  an  organism  to  a  single  host-plant  may  be  only  an  inference  based 
on  insufficient  observation  rather  than  an  actual  fact.  After  a  time  the  apple  and 
quince  were  added  to  the  pear  as  host-plants,  and  now  we  may  add  also  the  plum 
and  the  loquat. 

PATHOGENIC  OR   NON-PATHOGENIC  TO   ANIMALS? 

If  the  organism  will  not  grow  in  the  thermostat  at  37°  C,  or  grows  only 
feebly,  as  is  the  case  with  many  plant  parasites,  it  may  be  assumed  to  be  non-patho- 
genic to  animals  with  warm  blood.  Only  those  organisms  which  grow  readily  in 
the  thermostat  at  37°  C.,  and  which  closely  resemble  animal-pathogenic  forms  or 
which  are  suspected  of  causing  some  particular  disease  of  animals,  need  be  tested 


Fig.  72* 

by  animal  experimentation  for  economic  purposes.  In  general,  it  is  best  to  leave 
this  part  of  the  work  to  the  animal  pathologist,  for  the  same  reason  that  the  more 
abstruse  chemical  problems  are  turned  over  to  the  chemist. 

All  of  the  plant-parasitic  bacteria,  so  far  as  tested,  have  turned  out  to  be  non- 
pathogenic  to  warm-blooded  animals,  but  it  is  not  unlikely  that  some  exceptions 
may  be  discovered. 

Another  question,  of  special  interest  to  animal  pathologists,  arises  here,  namely, 
whether  forms  known  to  be  pathogenic  to  animals  and  especially  to  man  are  ever 

*Fic.  72. — Bacterium  pruni.  Vertical  section  through  a  green  plum  fruit  (var.  Hale)  showing 
bacterial  cavities  and  the  escape  of  the  organisms  through  the  ruptured  stoma.  In  this  case  beyond 
doubt  the  central  stoma  is  the  one  through  which  the  infection  originally  took  place.  Drawn  from 
a  photomicrograph.  The  material  was  fixed  in  alcohol,  infiltrated  with  paraffin,  cut  on  the  micro- 
tome, and  differentially  stained. 


PATHOGENIC   OR    NON-PATHOGENIC   TO   ANIMALS? 


harbored  by  plants.  Of  those  known  to  cause  animal  diseases  none  have  ever 
been  found  naturally  present  in  plants,  but  some  of  them,  such  as  the  typhoid 
bacillus,  the  anthrax  organism,  etc.,  have  been  shown  to  live  for  a  number  of 
days  or  weeks  when  injected  into  various  living  plants,  and  in  some  instances  have 
been  found  to  multiply  a  little  in  the  vicinity  of  the  wounds.  In  general,  their  life 
is  short  in  such  situations,  they  do  not  penetrate  far  into  the  tissues,  and  they  are 

manifestly  on  the  defensive.  If  they 
can  do  no  better  when  injected  into 
vegetable  tissues  in  enormous  quanti- 
ties, it  seems  rather  unlikely  that  under 
ordinary  natural  conditions  they  would 
find  their  way  into  plants  so  as  to 
make  them  dangerous  for  food.  In 
this  connection  the  reader  is  referred 
to  Volume  II,  where  this  subject  is 
discussed  more  fully.  More  danger  is 
likely  to  result  from  pathogenic  organ- 
isms carried  on  the  surface  of  plants, 
especially  on  salads  and  fruits  which 
are  not  cooked.  In  times  of  the  gen- 
eral prevalence  of  typhoid  fever,  chol- 
era, or  the  bubonic  plague,  the  writer 
for  one  would  certainly  prefer  to  forego 
salads  and  to  eat  only  freshly  cooked 
vegetables.  The  danger  from  such 
foods  in  time  of  epidemics  is  very 
great,  especially  in  localities  where 
ditch-water  is  frequently  sprinkled  on 
the  vegetables  to  freshen  them,  e.  g., 
in  parts  of  southern  Italy. 

Most  saprophytes  when  injected 
into  living  plants  behave  in  the  same 
way  as  the  animal  parasites,  i.  e.,  they 
either  die  at  once  or  maintain  a  pre- 
carious existence  for  some  weeks  in  the  vicinity  of  the  wound  and  then  succumb. 
The  writer  has  made  many  experiments,  with  negative  results.  The  most  extensive 
published  series  of  experiments  are  those  of  Zinsser  (Jahrb.  f.  wiss.  Bot.,  1897). 
To  get  a  particular  disease,  the  parasite  must  be  used  and  not  some  other  organism. 
This  the  writer  has  observed  over  and  over  again.  This  statement  holds  good  with 
plants  the  same  as  with  animals.  In  case,  however,  of  the  less  typical  plant  diseases 
(soft  rots)  various  members  of  a  group  of  closely  related  organisms  may  produce 
essentially  similar  phenomena.  This  is  paralleled,  however,  in  certain  of  the  less 
typical  animal  diseases. 

*Fic.  73. — Seedling  sweet-corn  plant  extruding  water  from  its  leaf-tips.  Most  of  the  infections 
by  Bacterium  Stewarti  take  place  during  this  stage  of  growth,  the  bacteria  passing  down  the  leaf 
through  its  vessels  and  entering  the  stem  through  the  lower  nodes.  Natural  size. 


Fig.  73.* 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


ECONOMIC   ASPECTS. 

The  economic   aspects  may  be   considered   under  four   heads:     (i)  Losses; 
(2)  Natural  methods  of  infection  ;  (3)  Conditions  favoring  the  spread  of  the  disease ; 

(4)  Methods  of  prevention. 

In  the  United  States  Department  of  Agriculture 
and  in  our  State  Experiment  Stations,  naturally,  much 
stress  is  laid  on  economic  considerations,  especially  on 
2,  3,  and  4.  A  knowledge  of  2  and  3  will  frequently 
lead  to  some  simple  and  effective  means  of  prevention. 

LoaSES. 

It  is  desirable  that  there  should  be  made  from  time 
to  time  a  careful  estimate  of  the  losses  caused  by  each 
particular  disease,  not  only  as  a  warning  to  farmers, 
fruit-growers,  market-gardeners,  and  florists  of  the  exist- 
ence of  these  dangers,  but  also  as  an  aid  to  legislatures 
and  governments  in  deciding  how  much  money  may 
be  judiciously  appropriated  for  the  scientific  investiga- 
tion of  these  problems.  Pathologists  are  urged  to  make 
and  publish  such  records.  It  is  perhaps  unnecessary 
to  add  that  the  determinations  should  be  reasonably 
accurate,  otherwise  it  were  much  better  not  to  make 
any  records.  Occasionally,  when  diseases  are  wide- 
spread and  destructive,  so  that  depreciation  of  land 
values  and  the  hostility  of  a  community  might  result 
from  great  publicity,  the  pathologist  may  have  to  con- 
sider discretion  the  better  part  of  valor  and  refrain 
from  publishing,  but  in  this  event  he  should  not  fail 
to  make  full  records  which  may  subsequently  be  pub- 
lished or  at  least  consulted.  What  we  need  and  must 
finally  have  is  a  large  body  of  accurate  statistics, 
covering  a  series  of  years,  many  localities,  and  many 
diseases.  To  make  these  statistics  most  useful,  certain 
meteorological  data  should  be  collected  in  the  same 
localities.  To  be  of  most  service  this  data  concerning 
the  weather  should  be  recorded  by  the  pathologist  him- 
self, who  will  be  better  able  than  anyone  else  to  note 
down  just  those  things  likely  to  influence  the  host- 
plants  favorably  or  unfavorably.  Some  of  these  things 


Fig.  74  * 


*Fic.  74. — Bacterium  Stewarti  (Erw.  Sm.)  attacking  sweet  corn  (Zea  mays).  The  section  was 
cut  from  the  extreme  upper  part  of  a  seedling  leaf  which  was  fixed  in  strong  alcohol  six  days  after 
placing  the  bacteria  on  its  tip.  At  the  time  of  inoculation  water  was  extruding  from  the  leaf-tip,  as 
shown  in  fig.  73.  This  figure  represents  a  longitudinal  vertical  cut.  The  dotted  and  heavily  shaded 
parts  show  the  location  of  the  bacteria  which  have  entered  through  the  ordinary  stomata  and  have 
not  yet  penetrated  the  vascular  system,  although  in  places,  as  at  D,  they  are  close  to  the  spiral  ves- 
sels. At  A,  B,  and  C  are  three  stomata.  The  substomatic  chamber  under  A  is  free.  B,  with  its 
surroundings,  is  shown  more  highly  magnified  in  fig.  75.  Drawn  with  help  of  the  Abbe  camera. 


COLLECTION    OF   STATISTICS. 


91 


are  cloudy  weather  (especially  if  prolonged),  sunny  weather,  frequent  or  excessive 
fogs  or  dews,  amount  of  rainfall,  and  frequency  of  rainfall,  snowfall,  hail,  excessively 
hot  weather,  cold  spells  and  frosts,  droughts,  daily  maximum  and  minimum  tem- 
perature, prevalence  of  special  diseases  correlated  with  special  peculiar  conditions, 
absence  of  other  diseases,  etc. 

NATURAL  METHODS  OF  INFECTION. 

Under  this  heading  the  student  should  be  on  the  watch  for  transmission  of  the 
disease  through  fungous  or  insect  injuries,  by  mollusks,  by  birds  or  quadrupeds, 
and  by  the  hand  of  man.  Man  contributes  to  the  spread  of  diseases  in  various  ways, 


Fig.  75* 

e.g.,  by  neglect  to  remove  diseased  plants,  by  use  of  infected  knives  and  other 
tools,  by  the  introduction  of  infected  seeds,  or  manures,  or  soils,  or  water,  and  by 
subjecting  his  plants  to  a  variety  of  depressing  and  unwholesome  conditions. 

A  great  variety  of  parasites  find  their  home  in  the  earth,  the  top  crust  of  which 
swarms  with  bacteria  and  fungi.  Such  parasites  are  frequently  introduced  from  one 
locality  to  another  in  infected  soils  adhering  to  wagons  and  other  farm  tools,  to  the 
feet  of  men  and  animals,  to  the  roots  of  transported  plants,  etc.  The  soil  is  a  living 
thing  and  it  should  not  be  transported  even  from  one  field  to  another  on  the  same 

*FiG.  75. — Bacterium  Stewarti  filling  the  substomatic  chamber  and  pushing  out  into  the  deeper 
tissues  of  a  maize  leaf.  The  result  of  an  inoculation  made  by  placing  a  small  quantity  of  a  pure 
culture  on  the  tip  of  a  sweet-corn  leaf  in  the  seedling  stage.  For  orientation  see  fig.  74.  The  glo- 
bose bodies  are  nuclei,  which  are  not  enlarged  (?). 


92  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

farm  without  due  consideration  of  what  may  happen.  Certain  bacterial  diseases 
might  be  distributed  very  readily  in  this  way  and  good  fields  rendered  worthless 
for  certain  crops. 

The  parasite  may  gain  entrance  to  the  plant  through  wounds  (plates  2  and  4 
and  fig.  8)  or  by  way  of  the  stomata  (figs.  70  to  75),  lenticels,  water-pores  (figs.  76 
to  79),  and  nectaries.  In  recent  years  the  writer  has  discovered  a  number  of  very 
characteristic  infections  by  way  of  the  stomata  and  the  water-pores,  which  are  only 
modified  stomata,  e.  g,,  in  cabbage,  mustard,  plum,  bean,  soy-bean,  cotton  (fig.  80), 


Fig.  76.* 

pelargonium,  larkspur,  broomcorn,  sorghum,  maize,  cucumber,  etc.  Pear-blight 
affords  one  of  the  most  striking  examples  of  wholesale  infection  by  way  of  the  nec- 
taries. The  wilt  of  cucurbits  affords  an  equally  good  example  of  infection  through 
wounds — namely,  leaf-injuries  due  to  beetles. 


*Fic.  76. — Bacterium  campestre.  Section  of  a  cabbage  leaf  parallel  to  the  surface  and  near  the 
margin,  showing  the  result  of  infection  through  the  water-pores.  The  tissues  are  browned  and  de- 
stroyed. Immediately  under  the  leaf-serrature  a  cavity  has  formed  and  the  bacteria  have  begun  to 
penetrate  into  deeper  parts  of  the  leaf  by  way  of  the  spiral  vessels,  not  all  of  which  are  occupied. 
This  figure  is  slightly  diagrammatic,  but  only  to  the  extent  of  omitting  the  protoplasmic  contents 
of  the  parenchyma  cells  and  of  introducing  six  occupied  spiral  vessels  which  belong  to  the  next 
section  in  the  series.  No  spiral  vessels  are  visible  in  the  lower  part  of  the  section  because  the 
knife  passed  just  below  them.  Material  collected  on  Long  Island,  July  16,  1902,  and  fixed  in  strong 
alcohol.  The  spirals  here  shown  are  a  little  too  densely  occupied  by  the  bacteria  to  make  a  good 
drawing  under  the  oil-immersion  objective,  but  a  little  farther  in  (beyond  X)  they  are  less  abundant 
and  entirely  satisfactory  for  this  purpose. 


ECONOMIC   ASPECTS. 


93 


CONDITIONS  FAVORING  THE  SPREAD  OF  THE  DISEASE. 

The  conditions  favoring  the  spread  of  diseases  may  be  wholly  telluric,  such  as 
high  temperature,  unusual  drought,  cold  weather,  fogs,  heavy  dews,  and  excessive 

or  continuous  rainfall.  These  diseases  may  be  favored  by 
lack  of  natural  drainage,  or  may  be  brought  on  by  a 
variety  of  causes  which  are  largely  within  the  control  of 
the  grower,  such  as  selection  of  improper  varieties,  i.  e., 
very  susceptible  ones,  overcultivation,  storage  at  too  high 
temperatures  (in  case  of  cabbage  and  root  crops),  the  use 
of  infected  soils,  or  manures,  or  seeds,  or  plants,  and, 
especially  in  hot-houses,  by  the  mismanagement  of  water 
and  heat,  and  by  the  neglect  to  destroy  the  first  diseased 
plants  that  appear  and  such  transmitters  of  disease  as 
insects  and  slugs,  which  frequently  abound  in  hot-houses. 


Fig.  77.* 


METHODS  OF  PREVENTION. 


In  case  of  certain  diseases,  copper  fungicides  have  been  found  useful,  e.  g.,  in 
walnut  bacteriosis  and  some  of  the  leaf  spots,  but  in  general  we  know  as  yet  very 
little  about  bactericidal  treatments.  In  the 
early  stages  of  an  outbreak  some  of  these 
diseases  may  be  controlled  by  extirpation  of 
the  affected  parts,  or  by  the  removal  of  whole 
plants  as  soon  as  they  show  signs.  Also,  if 
possible,  the  common  carriers  of  infection 
should  be  eliminated.  Finally,  one  should  not 
forget  that  the  substitution  of  resistant  vari- 
eties for  susceptible  varieties  is  one  of  the 
most  hopeful  methods  for  disposing  of  certain 
of  these  vexatious  diseases.  Whenever  any- 
thing specially  noteworthy  has  been  discov- 
ered in  the  way  of  treatment  it  will  be  mentioned  under  each  particular  disease. 


Fig.  78.f 


*Fic.  77. — Bacterium  campestre  from  the  cavity  shown  in  fig.  76,  illustrating  water-pore  infec- 
tion of  the  cabbage.  X  2,000. 

tFic.  78. — Bacterium  campestre  occupying  a  spiral  vessel  in  a  cabbage  leaf  near  a  group  of 
infected  water-pores.  The  tissues  to  the  right  and  left  of  this  vessel,  and  also  above  and  below  it 
(slide  223  33,  18.6  by  9.7),  are  entirely  free  from  bacteria.  The  body  of  the  leaf  and  all  its  inner 
tissues  up  to  within  a  few  millimeters  of  the  leaf-tooth,  and  also  the  outer  surface  of  the  leaf  up  to 
the  water-pores,  are  sound.  On  the  contrary,  an  unbroken  bacterial  occupation  can  be  traced  from 
this  vessel  outward  to  the  water-pore  region.  The  bacteria  in  this  vessel  are  also  less  abundant 
than  in  those  nearer  to  the  group  of  water-pores,  «'.  e.,  its  occupation  is  of  more  recent  date.  Even 
if  there  were  no  other  evidence  of  infection  by  way  of  the  hydatodes  than  that  afforded  by  this 
vessel,  the  presence  of  the  bacteria  in  it  under  the  circumstances  mentioned  points  conclusively  to 
marginal  (water-pore)  infection  as  their  only  possible  source.  The  position  of  this  vessel  is  in  a 
small  vein  a  little  below  and  at  the  left  of  X  in  fig.  76.  Its  distance  from  the  left  margin  of  the 
bacterial  cavity  is  one  field  of  the  16  mm.  Zeiss  objective  with  the  12  comp.  ocular.  Its  distance 
from  the  sound  leaf  margin  is  two-thirds  the  diameter  of  such  a  field.  A  nucleus  is  shown  at  n. 


94 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


GENERAL  CONSIDERATIONS. 

LOCATION  OF  THE  LABORATORY. 

If  possible,  the  laboratory  should  be  in  a  clean  building  in  the  middle  of  a  green 
lawn.  If  it  must  be  in  a  crowded  and  dirty  city  it  should  be  on  an  upper  floor,  as 
far  removed  as  possible  from  the  dust  of  the  street  and  from  the  tramp  of  feet.  It 
ought  not  to  be  located  on  streets  filled  with  the  dust  of  heavy  traffic.  If  a  ground- 
floor  or  basement  room  in  a  dirty  locality  is  the  only  available  place,  then  the  air 
which  enters  the  room  should  be  filtered  through  absorbent  cotton.  A  south  front 
is  desirable  for  the  mounting  of  a  heliostat  and  for  other  photographic  purposes ; 
a  north  light  is  desirable  for  microscopic  use,  if  one  is  to  work  at  the  instrument 
continuously.  By  arranging  one's  time  according  to  the  position  of  the  sun,  the 
light  from  east  or  west  windows  may  be  used  to  advantage  five  or  six  hours  a  day, 
which  is  quite  long  enough  to  fatigue  ordinary  eyes.  The  writer  has  managed  to 

get  along  very  well  without 
north  light  for  the  last  ten 
years.  If  one  decides  to  use 
with  the  microscope  only  ar- 
tificial light,  such  as  that  of 
the  Welsbach  burner,  work- 
rooms for  this  purpose  may 
be  located  anywhere.  If  pos- 
sible, several  rooms  should 
be  secured  and  apportioned 
to  the  various  kinds  of  work, 
e.g.,  general  laboratory  rooms, 
chambers  for  special  workers, 
sterilization-room,  thermo- 
stat-room, cold-storage  and 
stock-culture  rooms,  storage 
rooms  for  chemicals,  small 
glass-inclosed  rooms  for  transfer  of  cultures,  photographic  rooms,  dark  rooms  for 
developing,  etc.  The  general  photographic  rooms  should  have  overhead  light  as 
well  as  side  light. 

EQUIPMENT  OF  THE  LABORATORY. 

Many  pieces  of  apparatus  may  be  procured  from  time  to  time,  as  the  exigencies 
of  the  work  demand  or  as  the  funds  will  permit.  Other  apparatus  must  be  provided 
on  the  start,  and  some  of  it  when  the  building  is  constructed  or  reconstructed. 


Fig.  79* 


*FiC.  79. — Small  portion  of  a  cabbage  leaf  from  Long  Island,  New  York,  showing  characteristic 
water-pore  infections  due  to  Bacterium  campestre.  The  blackened  veins  correspond  to  the  location 
of  the  bulk  of  the  bacteria  which  have  gained  entrance  to  the  vascular  system  of  the  leaf  by  way  of 
the  groups  of  water-pores  situated  on  the  serratures  of  the  leaf,  particularly  those  which  are  conspic- 
uously blackened.  Those  parts  of  the  leaf  where  only  the  larger  veins  are  shown  were  green  and 
normal  in  appearance.  Coll.  July  16,  1902.  Drawn  from  a  photograph. 


PLATE   10. 


fl 

1  ore 
S~  *> 


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5     C 

en 

03  T 

IS- 
II 

*    r> 

?  1 


6 


EQUIPMENT   OF   THE    LABORATORY. 


95 


There  should  be  hot-water  pipes,  cold-water  pipes,  steam  pipes,  a  steam 
bath,  gas-pipes,  compressed-air  pipes,  exhaust-air  pipes  (plate  10  and  fig.  81),  and 
electrical  wires  for  light  and  motive  force.  There  should  be  thermostats,  water- 
baths,  cooled  rooms,  ice-boxes,  steamers,  dry-ovens,  autoclaves,  a  distilled-water 
outfit,  an  alcohol-still  (by  which  waste  alcohol  may  be  recovered  or  absolute  alcohol 
prepared),  an  ether-still,  filters,  gas-generators,  gas-furnaces,  anaerobic  apparatus, 
the  very  best  microscopic  outfits  including  apochromatic  lenses,  photographic  and 
photomicrographic  appliances,  liquid-air  receptacles,  cylinders  of  compressed  carbon 
dioxide  and  oxygen,  microtomes,  paraffin  baths,  glassware,  balances,  chemicals,  and 
many  minor  pieces  of  apparatus. 


Fig.  80* 


*Fic.  80. — Angular  leaf-spot  of  cotton  in  which  stomatal  infections  appear  to  be  the  rule.  This 
leaf  represents  the  secondary  stage  of  a  natural  infection,  i.  e.,  the  spots  have  browned  and  shriveled, 
and  they  involve  the  entire  thickness  of  the  leaf.  In  an  earlier  stage  of  the  disease  the  spots  are 
limited  to  the  under  side  of  the  leaf  (mesophyll),  and  occur  in  the  form  of  small  water-soaked, 
uncollapsed  areas  surrounding  stomata,  under  which  nests  of  bacteria  occur.  These  spots  gradually 
deepen  so  as  to  involve  the  palisade  tissue,  and  then  they  become  visible  on  the  upper  surface  of  the 
leaf.  The  spots  are  not  yet  shriveled  or  browned,  but  if  the  leaf  is  held  up  and  viewed  by  trans- 
mitted light  they  appear  as  translucent  areas,  while  by  reflected  light  they  are  dull  and  wet-looking. 
A  little  later  they  present  the  appearance  shown  in  this  figure.  The  writer  has  obtained  all  stages  of 
this  disease  in  Washington  by  spraying  upon  the  plants  young  agar  cultures  of  Bacterium  malvace- 
arum  suspended  in  sterile  water. 


96 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


In  general,  the  working  capacity  of  a  laboratory  will  be  greatly  increased  by 
giving  the  director  a  stipulated  sum  of  money  per  annum  and  carte  blanche  to  buy 
laboratory  necessities  whenever  and  wherever  and  in  whatever  quantity  he  sees  fit, 
requiring  only  that  he  submit  vouchers ;  also  by  the  employment  of  a  number 
of  subordinate  assistants  of  special  fitness,  to  whom  may  be  assigned  much  of  the 
purely  mechanical  and  routine  work  of  the  laboratory,  such  as  the  proper  cleaning 
of  glassware,  the  making  of  ordinary  culture  media,  the  keeping  alive  of  stock 
cultures,  the  preparation  of  staining  media,  the  embedding,  cutting,  and  staining  of 
microtome  sections,  the  making  of  photographs,  the  indexing  of  literature,  etc. 

No  scientific  man  should  be  willing  to  trust  any 
piece  of  work  in  his  own  line  to  an  assistant  unless 
he  can  do  it  as  well  himself,  or  better,  but  when  it 
has  become  to  him  the  merest  routine,  his  time,  if 
worth  anything,  can  be  more  profitably  employed 
in  something  else.  In  most  American  laboratories 
which  the  writer  has  visited  there  is  a  woeful  lack  of 
intelligent  subordinate  assistance,  such,  for  example, 
as  that  furnished  by  the  German  "Diener"  and  the 
Malays  of  Java.  Every  assistant  can  not  hope  to  be- 
come at  once  an  independent  investigator,  although, 
if  competent,  his  work  should  always  be  shaped 
toward  this  desirable  end. 

A  good  library  should  be  within  easy  reach,  and 
as  a  suggestion  to  this  end  a  list  of  useful  books  and 
papers  is  appended  under  the  head  of  Bibliography 
of  General  Literature.  A  card  catalogue  of  current 
literature  is  also  very  useful  and  in  time  becomes 
invaluable  if  properly  made. 


Fig.  81.* 


CARE   OF   THE   LABORATORY. 


The  laboratory  should  be  a  clean  place.  Its  walls  should  be  of  such  material 
that  they  can  be  rinsed  or  wiped  down  occasionally.  The  floors,  doors,  tables, 
window-sashes,  etc.,  should  be  wiped  every  day,  every  other  day,  or  at  least  every 
third  day,  with  clean  cloths  wet  in  distilled  water,  boiled  water,  or  clean  lake  or 
artesian  water.  The  use  of  river  water,  swarming  as  it  does  very  frequently  with 
all  sorts  of  bacteria,  is  not  to  be  commended  for  cleaning  purposes,  and  brooms 
should  be  taboo.  No  one  should  enter  the  laboratory  who  has  not  business  there, 
and  order  and  quiet  should  prevail. 


*FiG.  81. — End  of  the  vacuum-pipe  on  laboratory  table.  The  gage  serves  to  show  the  degree  of 
exhaustion,  i.  e.,  whether  there  is  any  leak  in  the  piping  between  the  engine-room  and  the  labora- 
tory. The  two  rooms  should  be  connected  by  a  speaking-tube. 


CULTURE    MEDIA. 


PREPARATION   AND   CARE   OF   CULTURE   MEDIA. 


97 


Everything  should  be  carefully  weighed  or  measured.  Everything  should  be 
clean  as  possible  to  begin  with.  By  water  is  usually  meant  distilled  water,  and 
this  should  be  free  from  copper  or  other  gennicidal  metals  (see  Bolton,  Bibliog., 
XXXVIII).  Moore  &  Kellerman  have  shown  very  recently  that  the  Bacillus 
typhosus  is  destroyed  in  distilled  water  if  the  merest  trace  of  metallic  copper  is 
present.  Water  swarming  with  this  organism  was  sterilized  simply  by  standing  three 
hours  in  a  copper  vessel.  The  writer  found  the  count  of  Bacillus  tracheiphilus  reduced  „ 
over  30  per  cent  by  exposure  in  bouillon  in  block-tin  tubes  for  twenty-one  hours. 
Exposure  for  forty-eight  hours  gave  the  same  result,  i.  e.,  33  per  cent  destroyed.  A 
simple  glass  still  is  shown  in  fig.  82.  As  far  as  possible  the  chemicals  should  be 


Fig.  82* 

c.  p.,  and  in  many  cases  it  is  necessary  to  make  the  test  for  oneself,  no  matter  what 
the  reputation  of  the  firm  or  the  statement  on  the  label.  When  possible,  broken 
packages  should  be  avoided.  It  is  therefore  best  to  procure  most  chemicals  in 
several  small  packages  rather  than  in  one  large  one.  If  the  preparation  of  culture 
media  is  broken  off  before  its  completion,  by  nightfall  or  interruptions  of  any  kind, 
the  unsterilized  or  incompletely  sterilized  media  should  be  put  into  the  ice-box, 
especially  if  it  is  warm  weather.  Neglect  of  this  precaution  frequently  results  in  the 
spoiling  of  the  media.  In  steam  sterilization  one  should  begin  to  count  time  only 
after  the  thermometer  registers  100°  C.,  or  at  least  99°  C.  Those  who  live  in  high 


*Fic.  82. — Portion  of  a  work-table  showing  method  of  distilling  water  for  use  in  making  culture 
media.  The  flasks  should  be  insoluble  glass.  The  cold  hydrant  water  passes  through  the  condenser 
in  the  direction  of  the  arrow.  In  actual  use  the  upright  flask  and  the  flame  are  sheltered  from  air- 
drafts  by  sheet  asbestos.  One-ninth  actual  size. 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


mountain  regions  must  use  autoclaves.  Agar,  potato,  etc.,  in  test-tubes,  may  be 
steamed  twenty  minutes  on  each  of  three  consecutive  days.  Gelatin,  beef-bouillon, 
and  all  other  fluids  likely  to  be  injured  by  long  heating  should  be  steamed  only  ten  or 
fifteen  minutes  on  each  of  three  consecutive  days,  if  in  tubes.  The  writer  frequently 
steams  such  media  fifteen  minutes  the  first  day,  ten  minutes  the  second,  and  five 
minutes  the  third.  Agar,  gelatin,  bouillon,  etc.,  stored  in  flasks  in  large  quantity 
must  be  steamed  a  longer  time — usually  thirty  to  forty-five  minutes  on  each  day. 

The  first  steaming,  when  softened  gelatin  is  added 
to  bouillon,  usually  requires  thirty  minutes.  To 
melt  flasked  agar  quickly,  shake  it  into  fragments 
or  break  it  with  a  sterile  glass  rod  before  putting 
it  into  the  steamer. 

Oversteaming  should  be  carefully  avoided.  It 
softens  gelatins  or  altogether  prevents  their  solidi- 
fication, and  is  very  apt  to  cause  troublesome  pre- 
cipitates in  a  variety  of  media.  Precipitates  in 
bouillon  often  occur  if  the  tubes  are  not  clean,  or 
if  the  bouillon  was  not  well  boiled  at  first  before 
filtering  and  placing  in  tubes.  If  the  beef-broth 
looks  greenish  in  the  beaker  or  flask,  rather  than 
a  clear  yellow,  it  may  be  assumed  that  it  needs 
more  boiling  and  that  if  tubed  in  this  condition  it 
will  throw  down  whitish  particles  on  subsequent 
steaming.  The  writer  prefers  to  obtain  his  ordi- 
nary +  bouillons  by  incomplete  neutralization 
with  sodium  hydrate  rather  than  by  addition  of 
hydrochloric  acid  after  full  neutralization.  The 
adding  of  hydrochloric  acid  precipitates  out  certain 
nutrient  substances  and  also  seems  to  interfere 
with  the  growth  of  some  organisms.  Distilled 
water  and  river  water  should  be  sterilized  in 
quantity  in  the  autoclave.  For  details  concern- 
ing the  making  of  particular  media  the  student 
should  consult  the  standard  text-books,  a  dozen  or  more  of  which  should  be  kept 
within  easy  reach  in  every  laboratory.  Some  formulse  are  given  in  the  middle 
part  of  this  volume.  The  autoclave  may  be  used  for  the  preparation  of  sterile 
water  and  some  media,  but,  in  general,  I  prefer  media  which  has  not  been  heated 
above  100°  C.,  especially  for  use  with  sensitive  organisms.  Media  should  be 
heated  in  the  autoclave  only  for  a  brief  time  and  at  a  minimum  pressure,  generally 
not  more  than  ten  minutes  and  at  not  more  than  110°  C.  Milk,  gelatin,  and 
media  containing  sugars  should  never  be  sterilized  in  the  autoclave.  Sugars 


Fig.  83  * 


*Fic.  83. — Apparatus  for  rapidly  filling  test-tubes  with  10  cc.  portions  of  agar,  bouillon,  etc.  By 
means  of  tins  device  an  expert  assistant  can  fill  500  tubes  an  hour.  Made  to  order  by  Emil  Greiner. 
Height,  23  inches.  The  bulb  above  X  is  essential. 


PREPARATION    AND    CARE   OF    CULTURE    MEDIA. 


99 


and  other  substances  decompose  at  these  high  temperatures  and  the  results 
obtained  by  the  growth  of  bacteria  in  such  media  are  not  comparable  with  those 
obtained  on  media  sterilized  at  100°  C.  Kitchens  has  recently  shown  that  detri- 
mental acids  are  formed  when  bouillon  containing  sugar  is  autoclaved.  Peptone 
water,  agar,  and  bouillon  may  be  sterilized  in  the  autoclave.  For  titrating  culture 
media  the  writer  uses  the  burettes  shown  in  fig.  59.  The  twentieth-normal  alkali 
is  stored  as  shown  in  fig.  60.  Quadruple-normal  sodium  hydrate  solution  is  used 
for  neutralization.  The  phenolphthalein  solution  is  made  by  adding  i  gram  of 

N 
the  dry  powder  to  100  cc.  of  50  per  cent  alcohol,  and  then  enough  —  sodium  hydrate 

to  carry  it  fully  into  solution,  removing  the  yellow  color  without  making  the  fluid 

a  very  decided  pink.  Fluid  media  may 
be  filled  into  tubes  very  rapidly  by  means 
of  the  device  shown  in  fig.  83.  For  storing 
media  sterilized  in  test-tubes  and  for  hold- 
ing cultures  made  on  such  media  the  writer 
has  found  ordinary  quinine  cans  very  use- 
ful  (fig.  84). 

The  proper  care  of  culture  media  after 
sterilization  involves  considerable  thought 
if  they  are  not  to  be  used  immediately. 
Stored  media  lose  water  and  along  with 
this  loss,  of  course,  there  are  physical 
changes,  so  that  the  results  obtained 
are  not  always  comparable  with  those 
obtained  from  similar  media  containing 
the  standard  volume  of  water.  Various 
devices  have  been  recommended  for  pre- 
venting this  loss  of  water.  Rubber  caps 
keep  in  the  moisture,  but  are  apt  to 
favor  the  development  of  fungi.  Paraf- 
fined  plugs  made  by  removing  the  cotton 
plug,  dipping  the  lower  end  of  it  quickly  into  and  out  of  hot  sterile  paraffin,  and 
replacing  it  in  the  mouth  of  the  tube  or  flask  before  the  melted  paraffin  has  had 
time  to  cool,  answer  the  purpose  very  well,  but  have  the  objection  that  all  of  the 
tubes  must  be  placed  in  turpentine  or  some  other  solvent  of  paraffin  before  they 
can  be  cleaned  for  a  second  use.  On  the  whole,  the  use  of  moderately  tight  plugs 
and  the  storage  of  the  media  in  cool  or  cold  air  are  the  best  methods  of  retaining 
the  water  content  of  the  medium.  Nutrient  media  should  be  made  in  small  quan- 
tities and  often,  rather  than  in  large  quantities  and  at  infrequent  intervals.  The 
cotton  should  be  dry-heated  in  bulk  before  plugs  are  made  from  it 


*Fic.  84. — Ordinary  quinine  cans  with  a  little  cotton   in   the  bottom   are   very   convenient   for 
holding  cultures  and  culture-media  in  test-tubes.    One-third  actual  size. 


100 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


THE   CLEANING   AND   STERILIZATION   OF   GLASSWARE   AND   INSTRUMENTS. 

New  glassware  may  be  boiled  in  soap-suds,  rinsed  thoroughly,  soaked  in  the 
chromic-acid  cleaning  mixture  for  some  hours,  rinsed  in  hydrant  water,  soaked  in 
several  changes  of  distilled  water,  soaked  or  shaken  in  alcohol,  and  finally  rinsed 
in  distilled  water.  Neglect  to  wash  in  alcohol  will  frequently  leave  behind  on  the 
walls  of  the  test-tubes  an  invisible  film  which  causes  vexatious  precipitates  in  beef- 
bouillon,  etc.  Discarded  tubes,  flasks,  and  dishes  containing  living  organisms  must 
be  autoclaved  or  filled  with  the  chromic-acid  cleaning  mixture  before  they  are  washed. 
Some  responsible  person  should  attend  to  this.  If  acid  is  used'it  should  be  allowed 
to  act  for  some  hours. 

Petri  dishes  should  fit  together  well,  but  not  tightly,  and  should  be  double- 
wrapped  in  clean  Manila  paper  before  placing  them  in  the  hot-air  oven,  or  else 
should  be  inclosed  in  suitable  tin  boxes.  The  writer  prefers  to  wrap  them.  The 
paper  for  this  purpose  may  be  12  by  12  inches.  The  dish  should  be  placed  in  the 

middle.  The  sides  of  the  paper 
are  folded  over  it;  the  corners 
of  the  projecting  ends  are  then 
turned  in,  leaving  V-shaped 
flaps,  which  are  folded  down  on 
to  the  plate.  The  second  cover- 
ing is  folded  at  right  angles  to 
the  first  and  on  the  other  side 
of  the  dish.  Dishes  treated  in 
this  way  and  ready  for  steril- 
ization are  shown  in  fig.  85. 
Pipettes  should  be  dry-heated 
in  the  tin  boxes  already  men- 
tioned (fig.  37)  after  having  the 
upper  end  carefully  plugged 
with  cotton,  which  should  not 
project.  Knives,  scalpels,  scrapers,  spatulas,  needles,  forceps,  etc.,  may  be  sterilized 
in  the  Bunsen  flame,  or,  if  needed  cold  in  quantity,  may  be  wrapped  in  Manila  paper 
or  put  uncovered  into  short  tin  boxes  and  heated  in  the  dry  oven  at  140°  C.  for  two 
hours.  Petri  dishes,  test-tubes,  and  all  other  apparatus  wrapped  in  paper  and  put 
into  the  oven  for  sterilization  by  dry  heat  should  have  air  spaces  between  them,  i.  e., 
they  should  not  be  crowded  together  tightly,  and  the  recording  thermometer  should 
project  well  down  into  their  inidst.  The  investigator  should  test  the  behavior  of  his 
oven  when  full  and  empty.  Many  cheap  ovens  give  very  different  temperatures 
in  different  parts,  especially  if  filled  with  apparatus,  so  that  cotton  or  paper  may  be 
scorched  in  one  part  and  not  sterilized  in  another.  The  best  oven  known  to  the 
writer  is  that  made  by  L/autenschlager.  The  improved  form  of  the  L,autenschlager 
oven  shown  in  plate  6  does  not  require  watching  and  gives  a  uniform  temperature 

*Fic.  85. — Petri  dishes  wrapped  in  two  layers  of  Manila  paper  and  ready  to  be  dry  sterilized. 
They  are  set  on  edge  in  the  oven. 


Fig.  85* 


STERILIZATION. 


IOI 


in  all  parts.  It  also  furnishes  a  maximum  temperature  with  a  minimum  con- 
sumption of  gas,  hot  air  being  fed  to  the  flame.  The  apparatus  has  an  inner,  outer, 
and  middle  wall.  A  horizontal  iron  gas  pipe,  of  the  relative  size  shown  in  the  front 
of  the  picture,  passes  entirely  around  the  apparatus  at  the  bottom  between  the  outer 
and  middle  wall.  On  top  in  this  tube  are  many  small  openings  through  which 
gas  escapes  and  when  lighted  forms  so  many  small  Bunsen  flames.  Air  is  drawn 
in  at  first  and  mixed  with  the  gas  in  the  middle  open  part  of  the  feed 
pipe  in  front.  The  products  of  combustion  escape  through  the  chimney- 
on  top  of  the  oven.  There  are  pilot  lights,  so  that  the  apparatus  is  set 
going  easily.  The  result  of  this  arrangement  is  that  the  middle  wall 
becomes  heated  very  hot,  and  consequently  the  air  between  this  wall 
and  the  inner  wall  rises,  cool  air  entering  through  holes  in  the  bottom 
to  take  its  place.  There  is  thus  created  a  powerful  upward  mount  of 
hot  air.  This  enters  the  oven  through  several  hundred  holes  in  its 
ceiling,  is  forced  downward  and  escapes  through  as  many  holes  in  the 
floor.  From  this  place  the  hot  air  is  continually  crowded  sidewise  and 

backward  through  brass  tubes 
into  the  furnace  chamber  where 
it  serves  to  support  the  com- 
bustion. 

Unless  the  dry-oven  has  a  very 
uniform  temperature  through- 
out, so  that  there  is  no  danger  of 
scorching  the  cotton,  plugged 
test-tubes  should  be  tied  together 
loosely  and  stood  on  end,  cot- 
ton uppermost.  Petri  dishes 
(wrapped  in  paper  as  directed) 
may  be  set  on  edge.  If  the 
test-tubes  have  been  properly 
cleaned,  dry -heating  is  not 
necessary  for  such  as  are  to  hold 
steam-heated  media,  provided 
the  cotton  used  for  the  plugs  is 
dry-sterilized  in  advance.  The 
best  surgeon's  absorbent  cotton 


Fig.  66  * 


is  not  too  good  for  this  work.  It  should  be  unrolled  and  put  into  the  dry-oven  in  a 
loose  armful  and  heated  just  below  the  scorching  point  for  several  hours  (2  to  3  hours 
at  145°  C.  will  answer),  with  occasional  unfoldings  and  turnings  so  that  all  parts 
may  be  heated  uniformly.  It  is  now  taken  out,  re-rolled  and  put  away  in  clean 
paper  until  needed.  By  this  means  all  fungous  spores  lodged  in  it  are  destroyed  and 

*Fic.  86. — Dr.  George  Meyer's  hypodermic  syringe,  made  by  Lautenschlager.  Desirable  on  account 
of  perfect  workmanship,  and  because  it  is  easily  sterilized  without  injury.  This  size  holds  I  cc.  By 
twisting  the  button  of  the  piston  the  packing  at  the  other  end  is  tightened  or  loosened  at  will.  The 
separate  parts  are  enlarged  one-fourth. 


IO2 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


an  oil  is  driven  off  which  otherwise  would  be  deposited  as  a  whitish  distillate  on 
the  inside  of  the  test-tubes  near  the  plugs.  Hypodermic  syringes  may  be  sterilized 
by  boiling  in  distilled  water  if  the  contaminating  organism  is  non-sporiferous, 
or  by  soaking  twenty-four  hours  in  5  per  cent  carbolic-acid  water  or  lysol  water  and 

a  subsequent  soaking  and  boiling 
in  pure  water.  The  writer  prefers 
the  Meyer  syringe,  made  by  Lauten- 
schlager  (fig.  86).  Syringes  which 
allow  the  culture  media  to  ooze  out 
around  the  piston  whenever  any 
strong  pressure  is  exerted  are  danger- 
ous and  should  never  be  used  with 
infectious  material.  Those  which  do 
not  admit  light  or  allow  the  experi- 
menter to  see  how  much  fluid  has 
been  used  or  whether  air  is  present 
are  unsatisfactory.  In  case  of  many 
plants,  needle-pricks  are  more  satis- 
factory than  hypodermic  injections 
(pi.  4  and  figs.  8  and  88).  Needles 
are  sterilized  in  the  open  flame  as 
needed. 

When  conveniences  are  not  at 
hand,  as  on  long  trips  in  the  country, 
the  kitchen-oven  may  be  used  for 
sterilizing  glassware,  or  even  an  open 
flame  (alcohol  lamp),  and  agar  and 
gelatin  for  the  making  of  poured 
plates  may  be  melted  by  placing  the 
tubes  in  hot  water  in  a  tin  cup  or  tea 
kettle,  but,  in  general,  the  writer  has 
not  found  the  rooms  of  ordinary  farm 
houses  very  well  suited  for  research 
work.  Usually  they  are  too  dusty. 

Surgeon's  gauze  is  very  conve- 
nient for  laboratory  use,  for  coarse 
filters,  wipe-cloths,  etc. 

Fig.  87.* 


*Fic.  87. — Early  stage  in  the  infection  of  a  cabbage  leaf  by  Bacterium  camfestre;  a,  epidermal 
layer  on  the  apical  part  of  the  tooth  of  a  leaf,  showing  one  of  the  four  stomata  ( X  )  full  of  bacteria. 
For  the  condition  immediately  under  X  see  b,  which  was  drawn  from  the  third  section  in  series,  the 
intermediate  one  including  part  of  the  guard-cells.  'Slide  338,  Bi,  stained  with  carbol-fuchsin. 
Drawn  with  the  Abbe  camera,  3  mm.  Zeiss  apochromatic  objective  and  12  compensating  ocular. 
Material  collected  and  fixed  8  days  after  infection,  which  was  accomplished  by  atomizing  upon  the 
plant  water  containing  a  pure  culture  of  Bacterium  campestrc  grown  on  slant  agar.  When  collected 
many  of  the  serratures  had  begun  to  show  traces  of  the  brown  stain  which  invariably  appears  when 
this  organism  grows  in  cabbage.  The  plant  was  inclosed  in  the  cage  shown  in  fig.  95,  and  was  ex- 
truding fluid  from  its  water-pores  when  it  was  sprayed.  X  500. 


HOW    TO   AVOID    CONTAMINATIONS. 


103 


THE   MAKING   AND  TRANSFERENCE  OK   PURE   CULTURES. 

In  addition  to  what  has  been  said  under  Pathogenesis,  the  following  suggestions 
may  be  of  service  to  the  beginner. 

For  the  making  of  plate  cultures  and  for  the  transfer  of  organisms  from  one 
culture  medium  to  another,  select  a  still  day  and,  if  possible,  a  day  when  a  gentle  rain 
or  snow  is  falling.  This  offers  ideal  conditions,  since  the  earth  is  wet,  the  outside 
air  has  been  washed  free  from  dust,  and  there  is  no  wind  to  stir  up  dust  within  the 
laboratory.  A  strict  adherence  to  this  rule  is  sometimes  very  inconvenient  and  it  is 


Fig.  88.* 

not  meant  to  be  iron-clad.  It  is,  however,  of  immense  service  in  keeping  cultures 
free  from  contaminations,  and  those  who  propose  to  disregard  it  should  remember 
that  haste  in  the  beginning  of  an  experiment  often  leads  to  vexation  and  delay  in 
the  end,  especially  when  the  success  of  the  experiment  depends  absolutely  upon 
the  purity  of  the  culture. 

*Fic.  88. — Soft  rot  of  green  cucumbers  inoculated  by  needle-punctures  from  a  pure  culture  of 
Bacillus  carotovorus.  The  only  parts  not  softened  are  those  through  which  the  infected  needle  en- 
tered, i.  e.,  the  parts  rubbed  with  mercuric-chloride  water.  In  each  a  little  button  of  tissue  under 
the  disinfected  area  did  not  decay.  The  sound  fruit  at  the  right  was  punctured  at  the  same  time, 
but  with  a  sterile  needle.  The  cucumbers  had  been  removed  from  the  vine,  but  were  not  flabby. 
They  were  exposed  after  inoculation  to  the  ordinary  air  of  the  laboratory.  The  photograph  was 
made  on  the  seventh  day.  About  two-fifths  natural  size. 


104 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


When  ready  to  make  the  transfers  or  to  pour  the  plates,  close  the  windows, 
wipe  up  the  tables,  and  wet  down  the  floor,  window-sashes,  etc.,  with  distilled  water 
or  boiled  water,  and  reduce  the  air-currents  within  the  laboratory  to  a  minimum 
(especially  when  transfers  are  to  be  made  in  the  open  room)  by  keeping  the  doors 
shut  and  restricting  the  movements  of  all  persons  who  may  be  in  the  room.  It  is 
much  better  to  do  all  of  this  work  in  specially  constructed  small  rooms  (plate  n) 
than  under  hoods  (plate  12).  Hoods  are  open  only  in  front.  They  may  be  made  of 
any  convenient  size.  The  one  here  figured  is  is  32  by  39  by  20^  inches,  outside 
measurements.  When  one  is  far  from  laboratories  small  hoods  may  be  extempor- 
ized out  of  clean  paper,  or  cultures  may  be  poured  and  transfers  made  inside  of  a 
clean  pail  or  jar,  turned  down  on  its  side.  Any  method,  in  fact,  which  restricts 

the  movement  of  air  past  open  plates  and 
tubes  will  be  found  serviceable. 

The  work-shelf  of  the  room  shown 
in  plate  1 1  faces  a  window  as  wide  as  the 
room,  and  extending  from  the  level  of  the 
shelf  to  the  height  of  the  other  windows 
in  the  room.  This  window  faces  south 
and  is  only  6  feet  from  a  well-lighted  win- 
dow in  the  outer  wall  of  the  building.  The 
room  also  receives  bright  light  from  the 
west  side.  At  the  front  end  of  the  shelf 
are  a  Bunsen  burner  with  cut-off  flame,  a 
box  of  safety  matches,  a  box  of  rubber 
bands,  and  two  tumblers — one  for  burned 
matches  and  one  for  platinum  loops, 
needles,  forceps,  etc.  Immediately  under 
this  part  is  a  narrow  drawer  for  pencils, 
note  paper,  knives,  etc.  At  the  back  end 
are  a  few  wrapped  Petri  dishes,  a  nivella- 
tion  apparatus,  a  flask  of  sterile  water,  and 
a  crate  of  media.  Underneath  this  part  is 


Fig.  89.* 


a  second  shelf  3  inches  below  the  first,  where  Petri  dishes  and  tubes  containing  solid 
media  may  be  put  out  of  the  light  as  fast  as  inoculated.  The  size  of  this  room 
(inside  measurement)  is  4  by  4  by  10  feet,  and  it  is  large  enough.  No  provision  is 
made  for  ventilation,  because  air-currents  in  a  culture-room  are  very  objectionable. 
The  windows,  walls,  and  floor  are  wiped  up  with  distilled  water  before  making 
transfers.  Outside  is  a  bit  of  the  author's  private  laboratory.  At  the  right  is  the 
microtome  and  behind  it  on  the  wall  are  deep  and  shallow  drawers ;  69  is  for  bulk 
paraffin;  70  A,  B,  C,  D,  E,  are  cut  into  small  compartments  used  for  paraffin  blocks. 
The  very  shallow  drawers  are  for  ribbons  which  can  not  be  mounted  the  day  they 
are  cut ;  72  has  a  series  of  shelves  opening  on  the  south  side  and  is  used  to  hold 
photographic  printing  frames. 

*FiG.  89. — Pine  block  with  inch  holes,  convenient  for  holding  test-tube  cultures  during  exam- 
ination, or  tubes  of  media  which  are  to  be  inoculated.     A  good  size  is  95/2  by  3l/2  by  i^  inches. 


PLATE  11. 


The  author's  culture-room. 

At  the  left  hand  (back)  are  narrow  ihelvn  lor  culture-media,  pipette-boxo,  etc.    At  the  right  a  the  work-ihelf ,  covered  with  plate  gU». 


PREPARATION  OF  POURED  PLATES. 


105 


The  agar  may  be  poured  at  42°  C.  in  case  of  organisms  whose  thermal  death- 
point  is  known  to  be  high  (50°  C.  or  above).  For  all  others  it  must  be  cooled 
carefully  to  40°  C.  before  inoculating  for  poured  plates.  This  requires  five  or  six 
minutes  in  the  water  bath  at  40°  C.  Even  this  temperature  is  too  high  for  some 
organisms  and  then  gelatin  at  30°  C.  may  be  used.  When  ready  to  pour,  take  a 
clean  absorbent  cloth  and  carefully  wipe  all  water  from  the  outside  of  the  tube  (the 
lips  of  which  have  been  previously  flamed  gently  with  a  rotation  of  the  tube  on  its 
long  axis),  lift  the  cover  of  the  dish  only  as  much  as  is  necessary,  hold  the  cover  aver 
the  dish  (not  at  one  side),  pour  quickly  but  gently,  and  re-cover,  tilting  the  dish 
about  quickly  but  gently,  if  the  fluid  has  not  already  covered  the  bottom.  To  en- 
tirely cover  the  bottom  sometimes  requires  a  smart  little  jerk,  if  the  agar  is  not 
very  fluid.  The  student  must  learn  to  work  rapidly  and  dextrously,  then  there 
will  be  no  complaint  that  the  agar  has  solidified  before  the  plates  are  poured.  The 

plates  should  be  set  on  a  level  shelf  while  the  agar  or 
gelatin  is  hardening,  or,  if  the  colonies  per  square 
centimeter  are  to  be  determined,  a  nivelling  appa- 
ratus such  as  that  shown  in  fig.  66  must  be  used, 
and  the  dishes  should  have  flat  bottoms.  When 
plates  have  been  inoculated  too  abundantly  to  secure 
subcultures  from  single  colonies,  these  may  some- 
times be  obtained  from  the  traces  of  agar  or  gelatin 
left  in  the  tubes  from  which  the  plates  were  poured. 
With  this  end  in  view,  these  tubes  should  be  re- 
plugged and  laid  away,  for  a  few  days,  the  lips  and 
top  of  the  tube  which  were  wet  by  the  agar  or  gelatin 
being  first  heated  hot  in  the  flame,  care  being  exer- 
cised not  to  crack  the  tubes. 

All  tubes  containing  fluids  should  be  opened  and 
inoculated  in  a  position  as  nearly  horizontal  as  their 
contents  will  permit,  and  tubes  of  solid  media,  such  as 
agar,  may  be  held  level  or  inverted  for  inoculation.  A  convenient  block  for  holding 
test-tube  cultures  during  examination  is  shown  in  fig.  89.  It  is  usually  best  to 
flame  the  plugs  slightly  before  their  removal,  particularly  if  they  have  been  exposed 
to  the  air  for  some  days.  As  an  additional  precaution  the  transfers  should  be  made 
under  a  glass  hood,  or  in  a  special  culture-chamber.  If  sterilized  needles,  loops, 
knives,  forceps,  pipettes,  or  anything  else  designed  to  be  used  in  making  the  transfers 
have  accidentally  touched  anything  wJwtsoever,  they  are  presumably  contaminated 
and  must  be  rejected  or  reflamed.  Do  not  handle  the  lips  of  test-tubes  containing 
gelatin  or  agar  from  which  plates  are  to  be  poured.  Your  hands  may  be  con- 
taminated by  resistant  spores.  Take  hold  of  the  tubes  lower  down.  To  economize 
gas  and  avoid  heating  the  air  of  the  small  work-chamber  to  an  uncomfortable  degree, 
small,  cut-off,  constant-flame  burners  are  very  convenient  (fig.  90). 

*Fic.  90. — A  constant  Bunsen  burner  with  cut-off  flame.    Very  useful  for  the  laboratory  table 
and  the  culture  room.     About  two-fifths  actual  size. 


IO6  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

Plates,  tubes,  and  flasks  containing  pure  cultures  or  designed  for  inoculation 
should  never  be  opened  in  the  general  laboratory  on  a  windy  day  or  in  air  currents. 
Pour  two  uninoculated  agar  or  gelatin  plates  in  the  proper  way.  Keep  one  covered 
and  uncover  the  other  fora  few  moments  in  a  current  of  air,  i.  e.,  as  long  as  the  time 
required  to  make  a  plate  culture.  Then  keep  the  two  plates  together  and  com- 
pare from  time  to  time.  A  few  experiments  of  this  sort  will  convince  the  most 
skeptical  of  the  necessity  of  avoiding  drafts. 

The  person  and  clothing  of  the  experimenter  should  be  as  clean  and  free  from 
dust  as  possible.  White  duck  coats  are  very  convenient.  They  show  at  .once  when 
they  are  soiled  and  need  washing  and  ironing. 

Organisms  which  for  some  reason  may  be  difficult  to  obtain  in  ordinary  plate 
cultures  and  which  differ  markedly  from  their  associates  in  some  particular  way, 
e; £••>  by  more  rapid  growth,  by  indifference  to  heat,  to  acids,  to  thymol,  to  chloro- 
form, to  absence  of  air,  etc.,  or  which  can  use,  as  food,  substances  which  will  not 
support  the  growth  of  most  bacteria,  may  sometimes  be  isolated  very  readily  by 
providing  conditions  suited  to  their  growth  and  unsuited  to  that  of  the  bacteria  with 
which  they  are  mixed.  This  is  Winogradsky's  principle  of  elective  culture.  As  he 
defines  it,  this  is  a  culture  "  which  presents  conditions  favorable  only  to  a  single 
definite  function  or,  more  exactly,  to  a  function  as  strictly  limited  as  possible."  Such 


Fig.  91.* 

media  or  methods  are  exactly  the  opposite  of  universal.  Nutrient  starch  jelly  and 
nutrient  silica  jelly  are  good  examples  or  such  media.  Nutrient  fluids  rich  in  acid 
potassium  phosphate  or  destitute  of  nitrogen  are  additional  examples. 

Heat  is  often  an  excellent  means  of  separation.  Winogradsky  separated  his 
Clostridium  pasteurianum  from  all  but  two  of  the  contaminating  species  by  heating 
ten  minutes  at  75°  C.  (Archives  des  Sci.  Biol.,  Vol.  Ill,  p.  310).  The  isolation 
of  Streptococcus  (Leuconostoc)  mesenterioides  by  L,iesenberg  &  Zopf  and  of  Bacillus 
hortulanus  by  Sturgis  are  other  examples  of  separation  by  heat.  Omelianski's 
separation  of  his  hydrogen-cellulose  ferment  from  his  methane-cellulose  ferment  by 
exposure  of  the  recently  established  methane  ferment  to  75°  C.  for  fifteen  minutes 
is  another  good  example. 

THE  FINAL  DISPOSAL  OF  INFECTIOUS  MATERIAL^ 

Diseased  material  should  not  be  left  around  the  laboratory  any  longer  than  is 
necessary.  When  it  has  served  its  immediate  purpose  that  which  is  not  to  be  pre- 
served permanently  should  be  thrown  into  the  furnace.  Small  amounts  may  be 
sterilized  by  putting  into  beakers  or  jars  and  covering  with  cleaning  mixture  or 
equal  parts  of  crude  sulphuric  acid  and  water.  Crude  vegetable  and  animal  sub- 

*Fic.  91. — Instrument  for  making  puncture-inoculations.  It  consists  of  a  bone  handle  with  a 
metal-screw  socket,  into  which  a  sewing  needle  is  thrust.  The  needle  is  usually  of  small  size — a 
No.  8  or  10. 


PLATE  12. 


Work-table  with  movable  frame  of  wood  and  glass. 

Bacteriological  trantfen  may  be  nude  under  this  frame  in  the  open  room  it  windows  and  dooti  are  kept  doted. 


DISPOSAL   OF    INFECTIOUS    MATERIAL. 


107 


stances  likely  to  become  moldy  must  never  be  stored  in  refrigerators  designed  for 
pure  cultures.  The  open  ice-box  is  the  proper  place  for  such  substances,  and  they 
must  not  be  left  there  indefinitely.  Some  people  have  a  mania  for  collecting  every- 
thing and  then  keeping  it  a  long  time  without  making  any  use  of  it.  An  ice-box 
treated  in  this  way  soon  becomes  an  intolerable  nuisance. 

Discarded  plates,  tubes,  slides,  covers,  pipettes,  contaminated  litmus  paper,  etc., 
should  be  autoclaved,  or  covered  or  filled  with  cleaning  mixture,  or  dropped  into  it, 
as  the  case  may  be.  Deep,  narrow  glass  jars  or  long,  rectangular  enameled  pans  are 
necessary  for  the  pipettes.  Soiled  hands  may  be  disinfected  with  mercuric-chloride 
water  (1:1000),  which  should  always  be  on  hand  in  the  laboratory  in  quantity  prop- 
erly labeled.  Slight  wounds  should  be  washed  five  or 
ten  minutes  in  this  fluid.  Surfaces  of  floors,  tables,  etc. 
soiled  by  spilled  bacterial  cultures  should  be  covered 
immediately  with  mercuric-chloride  water  (1:1000)  and 
wiped  up  carefully  after  ten  or  fifteen  minutes  with 
distilled  water.  Spilled  cultures  of  molds  should  be 
soaked  in  mercuric  chloride  ( i :  1000)  for  at  least  an  hour 
before  wiping  up.  Neglect  of  these  simple  rules  means 
the  seeding  down  of  the  ice-boxes,  culture-chambers, 
and  the  general  laboratory  with  all  sorts  of  resistant 
mold  spores  and  bacteria.  An  abundance  of  cheap  car- 
bonate of  lime  should  be  kept  on  hand  for  the  prompt 
neutralization  of  spilled  acids.  A  mass  of  cotton  waste 
is  convenient  for  the  prompt  mopping  up  of  spilled 
fluids. 

All  contaminated  needles,  loops,  knives,  scissors, 
forceps,  etc.,  may  be  sterilized  in  the  open  flame. 
Instruments  which  are  too  valuable  to  be  flamed  may 
be  sterilized  in  carbolic  acid  (5  per  cent)  or  formal- 
dehyd  (5  per  cent)  or  lysol  (5  per  cent).  Never  put 
down  a  platinum  needle  or  loop  which  has  been  used 


Fig,  92* 


in  making  transfers  until  it  has  been  passed  carefully  its  whole  length  through  the 
flame.    Dissections  are  best  made  on  trays  which  can  be  easily  cleaned  and  sterilized. 


*Fic.  92.— Compressed-air  tank  and  spray-tube.  The  one  here  shown,  made  by  Boeckel,  Phila- 
delphia, is  nickel-plated  and  very  substantially  constructed.  It  is  filled  by  means  of  a  small  pump 
similar  to  a  bicycle  pump.  The  gage  registers  up  to  100  pounds  per  square  inch,  but  40  pounds 
pressure  is  ample.  The  bacterial  fluid  is  placed  in  atomizers  of  the  form  shown  in  fig.  93.  The 
method  of  attachment  is  not  satisfactory.  This  device  is  very  convenient  when  trees  or  low  plants 
covering  a  considerable  area  are  to  be  inoculated.  Height,  29  inches.  The  same  firm  has  devised 
a  compact  traveling  outfit,  the  compressed-air  tank  being  about  one-half  the  size  of  the  one  here 
figured.  The  whole  is  packed  into  a  neat  portable  box,  and  the  only  disadvantage  is  the  small  size 
of  the  air-chamber,  which  requires  more  frequent  pumpings.  Of  course  the  apparatus  may  be  used 
equally  well  for  the  distribution  of  fluid  germicides  or  insecticides. 


loS 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


METHODS   OF   INOCULATION. 

Inoculations  may  be  by  punctures  with  a  delicate  needle  (fig.  91),  by  abrasions  of 
the  surface,  by  hypodermic  injection,  by  watering  the  soil  with  infective  material,  by 
plunging  aerial  parts  into  infectious  liquids  for  a  longer  or  shorter  time,  by  simply 
putting  the  bacteria  into  drops  of  water  on  parts  of  the  plant  and  protecting  from 
sunlight  and  evaporation  for  some  hours,  or  011  a  larger  scale  by  spraying  portions 
of  the  surface  with  very  dilute  culture  fluids  or,  preferably,  with  water  containing 
the  bacteria  (figs.  92,  93,  94),  by  brushing  or  rubbing  cultures  into  some  part  of  the 
surface,  by  allowing  insects,  snails,  etc.,  to  feed  on  diseased  material  and  then  colo- 
nizing them  on  healthy  plants.  The  writer  has  made  good  use  of  this  last  method  in 
case  of  three  different  bacterial  diseases.  Stomatal  infections  may  be  secured  by  sub- 
jecting the  plants  to  conditions  similar  to  those  occurring  in  nature  on  dewy  nights 
or  during  heavy  fogs  or  prolonged  rains,  i.  e.,  by  placing  the  potted  plants  on  wet 
sand,  atomizing  thoroughly  with  sterile  water  and  covering  with  tall,  roomy  bell-jars. 
The  experiment  should  be  undertaken  in  a  cool  rather  than  a  warm  house.  When 

the  right  conditions  have  been  obtained,  moisture 
covers  the  surface  of  the  plant  in  tiny  drops  which 
do  not  evaporate.  The  bell-jar  may  now  be  raised 
and  the  plant  again  atomized  lightly  with  steril- 
ized water  containing  the  bacterium.  The  best 
time  to  do  this  is  late  in  the  afternoon,  so  as  to 
take  advantage  of  the  cooler  night  temperature. 
When  the  bell-jar  is  returned,  which  should  be 
immediately  after  spraying,  it  should  be  covered 
with  cloth  or  paper  to  protect  from  the  light. 
Usually  bell-jars  should  be  removed  at  the  end 
of  twenty-four  hours,  but  exceptionally  they  may 
be  left  on  thirty-six  to  forty-eight  hours,  if  not 

exposed  to  the  sun.  Inoculation  cages  are  very  convenient  for  small  plants  (fig.  95). 
In  case  of  trees,  or  shrubs,  or  masses  of  tall  herbs,  tight-fitting  covers  of  tent-cloth 
will  be  found  serviceable  for  obtaining  conditions  similar  to  those  prevailing  in  wet 
weather.  They  may  be  left  on  i  to  3  days,  the  outside  of  the  tent  as  well  as  the 
plants  within  being  sprayed  with  water  often  enough  to  keep  everything  moist 
until  infections  have  been  secured. 

When  the  nature  of  the  plant  will  permit  it  and  when  only  a  few  inocula- 
tions are  to  be  made,  the  surface  which  is  to  be  punctured  should  be  rubbed  thor- 
oughly for  three  to  five  minutes  with  mercuric-chloride  water  (1:1000)  and  then 

*Fic.  93.— Atomizers  for  use  with  the  air- tank  (fig.  92).  These  are  made  by  the  Davidson  Rubber 
Company,  Boston,  Mass.  About  one-fourth  actual  size.  The  De  Vilbiss  sprayer,  made  in  Toledo, 
Ohio,  and  now  used  by  the  writer,  has  several  distinct  advantages.  It  is  all  metal  and  can  be  steril- 
ized in  boiling  water  without  becoming  twisted  out  of  shape,  it  can  be  attached  more  easily  to  large 
flasks  and  to  the  tube  leading  from  the  compressed-air  tank,  and  the  spray  may  be  directed  up,  down, 
or  straight  ahead  without  changing  nozzles.  It  requires,  however,  more  force  to  operate  than  the 
Davidson  sprayers,  and  consequently  is  less  convenient  when  used  with  a  hand-bulb. 


SURFACE    STERILIZATION.  IOC) 

washed  with  equal  care  in  sterile  distilled  water.  When  many  inoculations  are 
made  with  large  numbers  of  check  plants  and  when  due  care  has  been  taken  to 
work  under  conditions  such  that  accidental  contaminations  from  the  same  organ- 
isms are  not  to  be  feared,  the  writer  has  not  found  this  precaution  necessary.  The 
use  of  mercuric  chloride  should  be  avoided,  if  possible,  especially  on  leaves,  as  the 
writer's  experiments  have  shown  that  it  penetrates  into  the  plant  (some  plants)  for 
a  considerable  distance  and  prevents  the  action  of  the  bacteria  to  this  extent  (fig. 
88),  if  not  altogether,  as  has  happened  in  some  cases. 

THE   KEEPING   OF   RECORDS. 

If  one  contemplates  doing  much  work,  a  careful  record  of  what  has  been  done 
is  as  important  as  the  experiment  itself,  since  exact  remembrance  is  certain  to  pass 
away  with  lapse  of  time. 

In  all  his  work,  the  student  should  accustom  himself  to  make  very  exact 
statements,  so  that  others  may  be  able  to  follow  him.  For  example,  he  should 
not  describe  his  organism  as  "yellow"  or  "red"  without  qualifications,  since  there 
are  many  yellows  and  reds,  but  should  carefully  compare  it  with  some  standard 
color-scale  (Ridgway's,  Saccardo's,  Standard  Dictionary,  etc.),  and  govern  himself 

accordingly.  He  should  not  say, 
"  Organism  does  not  grow  at 
room-temperatures,"  but  rather 
should  state  the  temperature  at 
which  growth  does  not  occur,  as 
15°,  25°,  or  35°  C,  any  one  of 
which  may  be  "room-tempera- 
ture," depending  on  the  latitudCj 
altitude,  and  time  of  year.  He 
should  not  say,  "Organism  is 
killed  at  temperature  of  65°  C.," 

without  at  the  same  time  stating 
F"ic  94  * 

the  age  of  the  culture,  condi- 
tions of  exposure,  and  time  required,  which  might  be  ten  days  or  five  minutes. 

Every  independent  worker  will  in  the  end  devise  a  method  of  note-taking  which 
is  more  or  less  characteristic  of  his  personal  peculiarities  and  best  adapted  to  his 
own  particular  needs.  For  all  persons  there  is  no  one  best  method.  The  methods 
described  in  the  following  paragraphs  have  been  settled  upon  as  those  most  con- 
venient for  the  writer,  but  it  does  not  follow  that  they  are  the  most  economical  of 
time,  or  the  best  devisable,  or  the  ones  which  independent  workers  should  follow. 
They  are  here  given  as  hints  for  beginners  and  because  the  method  a  man  employs 
in  his  work  is  always  a  matter  of  more  or  less  interest  to  his  fellow-workers. 

First  of  all,  there  should  be  provided  a  record  book  in  which  the  method  of 
preparation  of  each  culture  medium  is  carefully  described.  This  should  be  a  good- 


*Fic.  94. — Hand-sprayer  which  may  be  used  for  distributing  bacteria  on  plants.     Some  form  is 
usually  kept  in  every  pharmacy  and  sold  as  a  cologne  atomizer. 


no 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


sized  book,  well  bound  in  leather,  so  as  to  stand  long  and  hard  usage.  The  entire 
quantity  of  a  culture  medium  is  known  as  a  "  stock  "  and  receives  a  special  number, 
which  is  written,  pasted,  or  stamped  on  any  flask  or  tube  that  contains  it  and 
which  serves  to  identify  it.  If  a  stock  is  subsequently  divided  and  a  portion  of  it 
is  treated  in  some  different  way,  e.  g.,  receives  more  sugar,  acid,  or  alkali,  this  por- 
tion receives  a  new  number,  or  the  old  number  with  the  addition  of  a  letter  of  the 
alphabet.  Each  stock  described  in  the  record  book  is  numbered  serially  from  i, 
and  the  book  continues  in  daily  use  as  long  as  the  laboratory,  or  until  it  is  filled 
with  records  and  carefulry  filed  away  as  "Culture  Media,  Volume  I." 

The  small  pocket  ledger,  No. 
492  of  A.  C.  McClurg  &  Co.,  Chi- 
cago, is  very  convenient  for  certain 
kinds  of  notes,  especially  those 
made  in  the  field  and  those  required 
for  the  identification  of  alcoholic 
specimens  and  stained  slides  (fig. 
112).  All  records  should  be  in 
ink,  of  a  sort  which  does  not  fade, 
and  in  field  work  a  good  fountain 
pen  is  invaluable.  Pencil  records, 
especially  those  made  with  rapid- 
writing  soft  pencils,  soon  become 
illegible  and  should  not  be  toler- 
ated except  on  paper  to  be  sub- 
jected to  steam  heat. 

Large  sheets  of  well-gummed 
paper  should  be  procured  and  the 
labels  cut  in  the  laboratory  to  the 
size  needed.  Labels  may  be  cut 
rapidly  in  quantity  with  the  appa- 
ratus used  to  trim  photographic 
prints  for  mounts.  When  exposed 
to  streaming  steam  such  labels 
come  off  easily,  and  it  is  best  not  to 
paste  them  on  the  tubes  or  flasks 


Fig.  95  * 


until  after  the  final  steam  steriliza- 


tion. In  moist  climates,  stock  quantities  of  such  gummed  labels  must  be  kept  in 
air-tight  boxes  or  between  sheets  of  paraffined  paper.  Test-tubes  in  crates  are  kept 
separate  during  steaming  by  writing  the  number  of  the  stock  on  a  slip  of  paper  and 
thrusting  this  into  the  crate  with  the  test-tubes.  The  number  should  be  written  with 
a  lead  pencil.  Faber's  pencils  for  writing  on  glass  are  useful  in  case  of  flasks  and 


*Fic.  95. — Small  cage  of  wood  and  glass  in  which  herbaceous  plants  may  be  placed  for  inocu- 
lation by  spraying.  The  inside  measurements  are  12  by  12  by  30  inches.  The  large  door  is  a  great 
convenience.  Hook-fastenings  are  better  than  spring  catches. 


RECORDS. 


Ill 


fermentation  tubes,  since  records  made  with  these  pencils  will  bear  streaming  steam. 
An  inexpensive  black  pencil  which  writes  on  clean  glass  very  readily  and  bears 
steam  well  (even  better  than  Faber's)  may  be  made  by  stirring  into  melted  beeswax 
enough  lamp-black  to  make  a  thick-flowing  liquid  (as  thick  as  will  flow).  This  is 

poured  into  molds  made  by  wrapping  writ- 
ing paper,  in  several  turns,  around  a  lead 
pencil  or  thick  glass  rod,  tying  near  one  end, 
removing  the  rod,  squeezing  the  other  end 
flat,  turning  over  its  edge,  and  fastening  this 
flattened  end  in  a  split  stick  or  clamp.  The 
paper  should  be  retained  as  a  cover,  the  string 
being  removed  and  the  loose  edge  pasted 
down.  A  dozen  such  pencils  may  be  made 
at  a  cost  of  10  cents.  In  the  absence  of  such 
pencils,  flasks  and  fermentation  tubes  may 
be  distinguished  in  the  steamer  by  dropping 
over  the  neck  different-sized  rubber  bands  or 
different  numbers  of  the  same  kind  of  band, 
or  by  writing  with  a  lead  pencil  the  number 
of  the  stock  on  a  square  of  letter  paper,  cut- 
ting a  hole  in  its  center  and  slipping  this 
over  the  neck  of  the  flask  or  tube.  When 
the  steaming  is  over,  the  regular  labels  should 
be  pasted  on  or  the  stock  number  written  on 
with  the  proper  pencil. 
All  plate  cultures  and  all  subcultures  made  on  a  given  day,  no  matter  of  what 
organism,  are  numbered  serially,  beginning  with  i.  These  are  i,  2,  3,  etc.,  of  that 
particular  day.  Those  of  any  other  day  are  also  numbered  i,  2,  3,  etc.  The  writer 


Fig.  %.* 


Fig.  97.* 

usually  numbers  his  plates  I,  II,  III,  etc.  Labels  may  be  pasted  on  the  covers  of  the 
Petri  dishes,  or  all  may  be  done  with  the  glass  pencil.  Cultures  in  tubes  subject  to 
frequent  handling  and  likely  to  be  needed  for  some  time  should  have  gummed-paper 
labels  written  in  ink.  The  above  transcripts  from  labels  on  four  test-tube  cultures 

*FiG.  96. — Labels  from  test-tube  cultures. 

*FiG.  97. — 'Wooden  labels  from  inoculated  plants. 


112 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


(fig.  96)  sufficiently  indicate  what  is  necessary  to  form  a  satisfactory  record.  This 
could,  of  course,  be  considerably  abbreviated  by  a  system  of  symbols  or  by  depend- 
ing to  a  larger  extent  on  the  "Notes." 

In  case  of  the  inoculations,  on  the  contrary,  only  as  many  series  are  made  use 
of  as  there  are  diseases  under  consideration.  Each  plant  is  generally  given  a  single 
number,  no  matter  in  how  many  places  it  may  be  inoculated,  the  separate  inocula- 
tions being  kept  distinct,  if  necessary,  by  sub-numbers.  Each  series  begins  with 


-$i*J^s.s&*&M  $^V/3f2bdC«&ty).  Ju^fi/s 


t  \ 


4,          (]    5-  IP  &<  • 

'.      //,  74  o  £ 

7    ' 


^    /^  * 


^-^f    Jo 


7;     ,/ro 


0 


"  ?• 

"    II , 


3,    3 


Fig.  98.* 

No.  i  and  continues  in  an  unbroken  sequence  as  long  as  the  disease  is  under  con- 
sideration. The  labels  written  on  soft  wood,  covered  for  this  purpose  on  one  side 
with  white  paint,  are  stuck  into  the  earth  or  wired  to  the  plant.  Transcripts  from 
two  such  labels  are  shown  in  fig.  97. 

*Fic.  98. — Three  sheets  showing  method  of  keeping  maximum  and  minimum  temperature  rec- 
ords.    One-half  actual  size. 


RECORDS. 


After  trying  various  methods,  the  writer  has  settled  down  (in  the  absence  of  a 
stenographer)  to  the  following  style  of  pen  and  ink  notes  on  cultures,  inoculated 
plants,  etc.,  as  extremely  flexible  and  convenient.  Reams  of  ordinary  typewriter 
paper  are  cut  crosswise  into  three  equal  portions,  so  as  to  form  slips  about  8  by  3^ 
inches.  As  many  of  these  as  are  necessary  for  the  particiilar  purpose  are  fastened 
together  at  one  corner  with  B,  J,  N,  C,  or  Z  eyelets  and  the  Triumph  punch,  sold 
by  The  W.  Schollhorn  Company,  New  Haven,  Conn.,  or  by  the  neat  little  saw- 


r~ 


Itf, 


S7W; 


72>. 


Fig.  99  * 

toothed  clamp  made  by  The  Middleton  P.  F.  Co.,  Philadelphia.  The  first  page  of 
the  slips  is  devoted  to  the  name  of  the  organism  under  examination,  the  kind  of 
experiment,  the  date  of  its  beginning,  etc.  The  subsequent  sheets  are  numbered 
serially  and  are  devoted  to  particular  plants  or  to  particular  cultures.  If  there  is  an 
overflow  in  any  particular  part  of  the  record,  it  is  very  easy  to  insert  additional 

*Fic.  99. — Sheets  showing  method  of  keeping  nitrate-bouillon  records.     One-half  actual  size. 


114 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


slips.  The  following  transcripts  from  actual  records  will  serve  to  illustrate  the 
method  (figs.  98  and  99).  As  fast  as  the  notes  are  completed  they  are  filed  away  in 
boxes  or  large  envelopes  until  the  whole  subject  has  been  worked  over,  when  they 
are  sorted  out  according  to  their  various  sub-heads,  and  all  the  data  which  they 

contain  is  thus  easily  available. 
The  writer  also  uses  a  sten- 
ographer whenever  possible, 
and  the  typewritten  sheets, 
after  immediate  careful  scru- 
tiny  for  errors  of  fact,  are  filed 
away  in  stout  Manila  envel- 
opes with  the  name  of  the 
parasite  written  on  one  corner; 


o6, 


fn- 


Fj 


16  by  12  inches  is  a  good  size 
for  the  envelopes. 

Card-catalogues  should  be  made  on  the  L.  B.  index  slips,  made  and  sold  by  the 
Library  Bureau,  Boston,  Mass.  Figure  100  is  a  sample  from  the  writer's  catalogue 
by  authors.  A  larger  size  should  be  selected  if  it  is  desired  to  include  abstracts. 
When  long  abstracts  or  considerable  extracts  are  made  from  literature  which  has 
been  borrowed,  or  may  not  be  readily  accessible  in  futitre,  heavy  sheets  (6^$  by  83^ 


32. 


XT    sS;O^^--7-^£^^_ 


(7  ... 


Fig.  101.  t 


inches)  have  been  used  by  the  writer.  These  have  headlines,  as  shown  in  fig.  101, 
and  are  preserved  by  tying  into  covers  made  for  the  purpose.  A  red  line  down  the 
left  side  of  the  sheet  preserves  a  space  for  a  marginal  index. 

A  serious  objection  to  the  making  of  many  abstracts  is  the  time  involved  and 
the  danger  of  degenerating  into  a  mere  student  of  literature  in  the  effort  to  make  a 
complete  catalogue  ;  another  is  the  fact  that,  if  made  in  advance  of  actual  need,  or 


*Fic.  100. — Sample  from  card-catalogue.    Two-thirds  actual  size. 

fFic.  101. — Top  of  large  sheet  used  for  voluminous  abstracts.  A  red  line  near  left-hand  mar- 
gin marks  off  a  space  on  which  summarizing  catch-words  or  phrases  are  written.  Breadth  of  sheet, 
6^  inches. 


RECORDS. 


"5 


• 


by  some  one  not  entirely  familiar  with  the  subject,  it  not  infrequently  happens  that 
the  statements  in  the  paper  which  have  been  omitted  from  the  abstract  as  unim- 
portant prove  in  the  end  to  be  the  essential  ones  so  far  as  the  owner  of  the  abstract 
is  concerned.  For  this  reason,  when  they  are  within  reach,  the  writer  prefers  to 
consult  the  original  papers  and  to  save  for  original  work  the  time  consumed  in 
making  long  abstracts.  When  they  are  rare,  frequently  needed,  and  only  to  be  had 

by  borrowing,  the  writer  has  sometimes 
photographed  the  more  essential  parts. 
In  one  instance  a  pamphlet  was  bor- 
rowed from  Europe  for  this  purpose. 

For  the  exact  measurement  of  col- 
onies, etc.,  a  strip  of  plate  glass  35  cm. 
long  and  ruled  into  350  mm.  spaces 
may  be  had  from  Carl  Zeiss,  and  will 
be  found  very  convenient  (fig.  102). 

Steel  rules  of  any  size  and  of  very 
excellent  workmanship,  graduated  ac- 
cording to  the  English  or  the  metric 
system  in  any  degree  of  fineness,  may 
be  had  from  the  L.  S.  Starrett  Com- 
pany, Athol,  Mass.  Two  of  these  rules 
much  used  by  the  writer  are,  respec- 
tively, 12  inches  and  30  centimeters 
long.  They  are  one  inch  wide  and 
about  three  sixty-fourths  of  an  inch 
thick.  They  are  graduated  on  both 
sides,  the  metric  rule  into  centimeters, 
millimeters,  and  one-half  millimeters, 
and  the  English  into  inches,  halves, 
quarters,  eighths,  sixteenths,  thirty- 
seconds,  and  sixty-fourths. 

Stage  micrometers  made  by  Zeiss 
are  recommended  for  the  finer  measure- 
ments. These  have  i  millimeter 
divided  into  tenths,  twentieths,  and 
one-hundredths  very  accurately.  All 
the  magnifications  of  microscopic 
objects  figured  in  this  book  are  recorded  in  terms  of  such  a  micrometer.  After  the 
drawing  has  been  made  it  is  customary  to  substitute  for  the  section-slide  this  stage 
microineter  and  throw  the  image  of  some  portion  of  the  ruled  scale  on  the  paper 

~ *FiG.  102.— Green  cucumber  soft-rotted  by  Bacillus  aroideae.  Contents  emptied  out  and  skin 
filled  with  water  and  so  photographed,  3  days  from  date  of  inoculation,  which  was  by  means  of  a 
few  needle-pricks.  The  fruit  was  kept  at  about  25°  C.  The  black  bands  are  pencil  marks  on  the 
millimeter  rule  placed  inside.  The  numerous  small  dark  spots  are  denser  bits  of  tissue  which  did 
not  wash  free  on  rinsing  out  the  sack  with  water.  At  the  left  drops  of  water  may  be  seen  oozing 
through  the  skin  and  falling.  Photograph,  nearly  natural  size,  by  Townsend. 


Fig.  102* 


I  ID  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

where  it  is  drawn,  taking  care,  of  course,  in  case  of  high  magnification,  to  start  one 
cross  line  from  the  outside  and  the  other  from  the  inside  of  the  image  of  the  lines. 
This  method  of  recording  magnifications  is  urged  on  all.  It  takes  but  a  moment, 
does  away  with  troublesome  computations,  and  enables  anyone  at  any  time  to  deter- 
mine just  what  was  the  magnification.  The  magnification  is  determined,  of  course, 
by  dividing  the  apparent  size  by  the  actual  portion  of  the  scale  shown.  For 


Fig.  103* 

example,  if  the  scale  drawn  on  the  paper  is  10  mm.  long  and  represents  o.oi  mm. 
of  the  actual  micrometer  scale,  then  the  magnification  is  X  1000;  if  it  represents 
the  entire  millimeter  of  the  micrometer  scale,  the  magnification  is  X  10. 
For  fine  weighings,  Christian  Becker's  balances  are  very  satisfactory. 


*Fic.  103. — Pillsbury  slide-boxes  empty  and  full,  made  by  Bausch  &  Lomb,  Rochester,  N.  Y. 
These  boxes  are  simple,  inexpensive,  and  satisfactory,  especially  for  serial  sections. 


COLLECTIONS. 


117 


THE   MAKING   OF   COLLECTIONS. 

A  good,  representative  collection  of  diseased  material  is  a  prime  necessity  in 
every  pathological  laboratory.  This  grows  into  completeness  only  with  the  lapse 
of  much  time  and  the  aid  of  many  hands.  It  should  include  photographs,  drawings, 
paintings,  dried  material,  representative  specimens  preserved  in  strong  alcohol,  and 
serial  sections  properly  stained  and  mounted  in  Canada  balsam  or  Dammar  balsam, 
which  must  not  be  dissolved  in  chloroform,  since  this  gradually  removes  the  stain. 
With  the  accumulation  of  much  material,  some  sort  of  classification  becomes  im- 
perative. At  present  the  writer  keeps 
the  material  designed  for  sections  in 
95  per  cent  alcohol,  arranged  in  as 
many  groups  as  there  are  parasites 
involved.  Each  jar  of  material  finally 
receives  the  same  number  as  the 
paraffin  block  from  which  sections 
are  cut.  This  material  must  be  exam- 
ined at  least  once  a  year  to  see  that 
the  alcohol  has  not  evaporated,  es- 
pecially if  corks  are  used.  Only  the 
best  velvet  corks  should  be  pur- 
chased, and  as  an  additional  precau- 
tion they  should  be  sealed  in  with 
paraffin.  The  negatives  are  filed  away 
in  similar  groups,  protected  by  nega- 
tive bags.  The  stained  sections, 
mounted  in  balsam,  are  filed  away 
in  cheap  wooden  boxes  (Pillsbury 
boxes),  each  holding  25  slides  (figs. 
103,  104).  These  are  very  conven- 
ient, if  properly  made,  but  some 
boxes  of  this  sort  lead  to  much  vexa- 
tion of  spirit,  the  grooves  being  too 
narrow  to  receive  any  but  the  thin- 


Fig.  104.* 


nest  slides.  Those  sold  in  recent  years  by  Bausch  &  L,omb  have  given  no  trouble. 
In  the  form  shown  in  fig.  104  the  cover  remains  on  better  and  the  mounted  slides 
are  easier  to  take  out,  but  in  drying  the  preparations  with  the  cover  off,  these  boxes 
tip  over  at  the  least  touch.  During  this  drying,  which  requires  from  a  few  days  to 
several  weeks,  the  slides  should,  of  course,  lie  flat,  not  on  edge. 

*Fic.  104.— Another  style  of  slide-box.  The  advantages  of  this  box  are  that  the  cover  is  not 
likely  to  fall  off  and  that  the  slides,  in  case  of  full  boxes,  are  withdrawn  more  easily.  The  disad- 
vantages are  that  it  is  tipped  over  very  easily  when  standing  on  end  open,  that  the  cover  is  readily 
mistaken  for  the  bottom  when  it  is  closed,  and  that  if  the  cover  is  put  on  upside  down  the  writing 
on  the  edges  is  divided.  These  may  also  be  had  from  Bausch  &  Lomb. 


n8 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


The  writer  passes  material  designed  for  sections  from  alcohol  through  chloroform 
(or  xylol)  into  paraffin.  Chloroform  is  preferred  in  case  the  infiltration  is  to  be 
completed  in  vacuo  ;  otherwise  xylol  is  generally  employed.  A  mixture  of  xylol 
and  alcohol  is  first  used,  then  pure  xylol,  after  this  xylol  with  as  much  paraffin  as 
can  be  dissolved  in  it  cold.  The  vial  is  then  placed  on  top  of  the  paraffin  bath  and 


Fig.  105* 

shaved  paraffin  added  until  it  will  dissolve  no  more  at  this  temperature ;  the  material 
is  then  placed  inside  the  apparatus  in  pure  melted  paraffin,  and  it  is  finally  mounted 
from  a  second  dish  of  pure  paraffin.  The  temperature  of  the  paraffin  bath  is  usually 

*Fic.  105. — A  small  paraffin  oven  much  used  in  the  writer's  laboratory.  The  capacity  of  the 
chamber  is  6  by  7  by  5  inches.  The  thermo-regulator  is  like  that  shown  in  fig.  35,  but  with  chloro- 
form substituted  for  glycerin. 


PARAFFIN-INFILTRATION. 

kept  at  59°  C.,  and  the  material  is  subjected  to  this  temperature  only  long  enough 
to  secure  proper  infiltration.  Generally  a  few  hours  are  sufficient.  A  small  oven 
used  for  this  purpose  is  shown  in  fig.  105.  For  large  laboratories  or  classes  of 
students  the  separate-compartment  paraffin  oven  designed  by  Dr.  Lillie  is  very 
convenient.  Griibler's  paraffin  is  preferred,  and  for  the  climate  of  Washington  we 
use  mixtures  of  three  grades  of  hardness,  viz,  melting  point  52°  C.,  58°  C.,  and 
60°  C.,  increasing  or  decreasing  the  amount  of  the  harder  sorts  according  to  the 
time  of  year.  Dirty  paraffin  should  never  be  used.  All  the  stock  paraffin  should 

be  carefully  protected  from  dust.  The  same 
remark  applies  still  more  pertinently  to  the 
sections  cut  on  the  microtome.  They  should 
be  made  in  still  air,  in  a  clean  room,  and  should 
be  carefully  protected  from  dust  until  stained 
and  mounted.  The  paraffin  -  infiltration  is 
usually  a  simple  process  unless  the  material 
contains  air.  The  embedded  material  is  given  a 
serial  number  which  is  scratched  on  the  paraffin 
(fig.  106),  until  it  is  fastened  to  the  cutting 
block,  when  it  is  written  on  the  latter  (fig. 
107).  These  blocks  are  kept  as  shown  in  fig. 
1 08.  The  sections  are  fastened  to  clean  slides 
*  by  a  very  thin  layer  of  Mayer's  egg  albumen 

fixative  (see  Lee's  Vade  Mecum,  5th  ed.,  p.  143),  or  with  pure  water,  or  preferably 
with  0.5  per  cent  gelatin  water  (which  will  not  keep  untreated,  but  may  be  preserved 
by  adding  3  per  cent  phenol) ;  the  paraffin  is  removed  (after  cautious  melting)  by 
exposure  to  turpentine  or  xylol,  alcohol  is  then  substituted,  and  thereafter  graded 
mixtures  of  alcohol  and  water  down  to  alcohol  containing  50  or  60  per  cent  of 
water,  followed  by  the  stain.  Water  is  then  removed  by  passing  through  graded 
alcohols  into  absolute  alcohol ;  xylol  or  bergamot  oil  is  substituted  for  the  alcohol, 
and  the  section  is  finally  mounted  in  balsam.  Coplin's  staining  jar  is  preferred 
(figs.  109,  no).  A  series  of  staining  jars,  ready  for 
use,  is  shown  in  fig.  in.  The  section  properly  fast- 
ened to  the  slide,  and  dry,  is  started  in  at  the  left  after 
melting  the  paraffin  with  gentle  heat,  and  is  taken  out 
at  the  right  ready  for  mounting  in  balsam.t  In  this 
series  of  jars  the  gradations  are  as  follows,  beginning 


*Fic.  106. — Infiltrated  tissues  embedded  in  paraffin  in  a  watch-glass  and  now  ready  to  cut  out 
and  mount  on  blocks  for  the  machine. 

fFic.  107. — Infiltrated  material  embedded  in  paraffin  and  mounted  on  a  pine  block  ready  to  cut 
on  the  microtome.  Actual  size. 

J  Sections  designed  for  photo-micrographic  work  must  not  only  be  cut  in  clean  air,  but  mounted 
in  absolutely  clean  balsam.  So  much  trouble  has  been  experienced  in  finding  such  dissolved  bal- 
sam on  the  market  that  the  writer  now  makes  his  own.  The  dry  balsam  is  heated  in  an  oven  until 
all  easily  volatile  products  are  driven  off  and  it  becomes  brittle.  It  is  then  dissolved  in  xylol  and 
filtered  under  a  bell  jar  to  exclude  dust.  The  filtering  usually  requires  several  days. 


I2O 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


I 


at  the  left  :  Xylol,  second  xylol,  xylol  one-third  absolute  alcohol  two-thirds,  95  per 
cent  alcohol,  75  per  cent  alcohol,  55  per  cent  alcohol,  40  per  cent  alcohol,  carbol- 
fuchsin,  40  per  cent  alcohol,  second  40  per  cent  alcohol,  55  per  cent  alcohol,  65  per 
cent  alcohol,  75  per  cent  alcohol,  95  per  cent  alcohol,  absolute  alcohol,  second 
absolute  alcohol,  xylol,  second  xylol.  From  this  last  jar  the  material  is  mounted 
in  balsam.  Turpentine  may  be  substituted  for  xylol  in  jars  i  and  2.  After  the 
paraffin  is  fully  removed,  the  slides  are  passed  rapidly  from  jar  to  jar  (a  minute  or  two 

in  each  being  generally 
sufficient)  until  the  stain 
is  reached.  After  remain- 
ing in  the  stain  the  proper 
length  of  time  (usually 
three  to  ten  minutes,  but 
sometimes  much  longer) 
the  slides  usually  are 
allowed  to  remain  in  the 
40  per  cent  alcohols  for  a 
number  of  minutes,  with 
frequent  inspection.  When 
they  appear  to  be  properly 
bleached  (rather  pale)  they 
are  passed  rapidly  through 
the  remaining  jars  until 
they  reach  the  xylol,  in 
which  they  may  remain 
for  some  time  without 
injury,  if  they  can  not  be 
mounted  immediately,  but 
they  must  not  be  allowed 
to  stand  for  any  great 
length  of  time  in  any  of 
the  alcohols.  The  secret 
of  success  lies  in  obtain- 
ing just  the  proper  amount 
of  differentiation  in  the 
40  per  cent  alcohol  and  in 
not  losing  any  of  this  later 
on.  To  retain  the  stain  it 


Fig.  108* 


is  necessary  sometimes  to  omit  some  of  the  graded  alcohols. 

The  time  required  for  properly  staining  sections  varies  from  one  or  two  minutes 
to  a  half  day  or  more,  according  to  the  subject  and  the  stain  employed.  No  general 
rule  applicable  to  all  cases  can  be  given.  When  the  material  is  selected  for  embed- 
ding, its  serial  number,  with  a  full  description,  is  entered  in  the  record  book  (fig.  112). 


*Fic.  108. — One  of  a  series  of  drawers  divided  into  small  compartments  for  holding  infiltrated, 
embedded  material,  cut  and  uncut. 


PLATE  13. 


•n 
-i 
o 

3 

ri- 
IJ 

I"  ^ 
I  ? 
f  » 

3.     2. 


S    S 

I'  6 


RECORDS. 


121 


This  book  must  not  be  lost  or  misplaced.  The  advantage  of  having  the  serial 
number  written  also  on  the  bottle  containing  the  stock  of  preserved  material  is  very 
evident  if  a  thing  of  this  sort  ever  happens.  The  serial  number  is  written  on  one 

edge  of  the  slide-box,  and  serves  to  identify  it  (fig.  103).  

Some  record  besides  a  mere  number  should  also  be  placed 
on  the  slide-boxes.  All  the  slides  within  bear  this  num- 
ber, e.  g.,  256,  and  also  a 
series  number  of  their  own, 
i.  e.,  i  to  25.  The  slide- 
boxes  are  then  filed  away 
on  shelves  either  serially 
or  in  groups,  according  to 

the  parasite.     Slides  con-     N "** 

taining  particularly  good 

fields  are  marked  X,  and  when  the  best  fields  are 
finally  decided  upon  their  location  is  recorded  as  de- 
termined on  the  mechanical  stage.  In  case  a  dozen 
or  more  serial  sections  are  included  on  one  slide  the 
the  extra  good  ones  are  marked  X  on  the  first  exam- 
ination, and  the  others  O,  as  shown  in  fig.  113. 
When  one  of  these  sections  has  been  drawn  or  photo- 
graphed, the  X  is  underscored  or  inclosed  by  a  circle. 
This  method  enables  one  to  keep  track  of  any  num- 


Fig.  109* 


ber  of  sections.  Free-hand  sections  may  be  made  with  the  Torrey  razor  shown  in 
fig.  ii4D.  This  is  altogether  the  best  razor  the  writer  has  used.  When  very 
dull  it  may  be  sharpened  on  an  India  oil-stone.  These  stones  are  said  to  be  made 
of  a  mixture  of  carborundum  and  clay,  baked  at  a  high  temperature.  They  may  be 


Fig.  1114 

had  of  the  Norton  Emery  Wheel  Company,  Worcester,  Mass.,  in  three  grades  of  fine- 
ness, the  finest  being  usually  coarse  enough  for  the  dullest  razors.  The  size  needed 
is  8  by  2  by  i  inch.  The  finishing  may  be  done  on  an  Arkansas  oil-stone,  with  a 

*Fic.  109. — Coplin's  staining  jar.     About  one-half  actual  size, 

f  FIG.  no. — Cross-section  of  Coplin's  staining  jar.    About  actual  size. 

JFic.  in. — A  series  of  Coplin's  jars  filled  and  properly  arranged  for  staining  sections  fastened 
to  slides. 


122 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


few  final  touches  on  a  good  leather  strop.     The  maintenance  of  good  edges  on 

microtome  knives  is  a  matter  of  great  importance  and  considerable  difficulty,  and 

where  much  material  is  to  be  cut  it  is  very  economical  of  time  to  send  away  such 

knives  to  be  put  in  order  by  some 
expert.  In  recent  years  the  writer 
has  sent  all  such  knives  to  Charles 
L/entz  &  Sons,  Philadelphia,  with 
very  satisfactory  results.  Knives 
suitable  for  serial  sections  are  shown 
in  fig.  1 14  A  and  C.  In  fig.  1146  is 
shown  one  of  a  set  of  knives  not  in- 
clined to  spring  and  well  adapted  to 
the  cutting  of  hard  material  with  a 
long  slant  stroke.  These  knives  were 
made  to  order  by  L/entz  &  Sons  at  a 
cost  of  about  $6  each.  An  end-on 
view  of  all  these  knives  is  shown  in 
fig.  1 14  a,  &,  c,  d.. 

Many  plant  tissues,  especially  ma- 
ture leaves,  are  full  of  very  hard  cal- 
cium oxalate  crystals,  and  the  difficul- 
ties of  properly  cutting  such  material 
are  very  great.  The  cutting  of  thin 
sections  of  bone  would  be  quite  as 
easy.  After  even  a  few  sections  the 
edge  of  the  knife  looks  like  a  minia- 
ture saw  and  the  sections  themselves 
are  badly  torn,  partly  by  the  dulled 
knife  and  partly  by  the  movement  of 
p.  |j2*  the  crystals  themselves.  In  case  of 

the  yellow  disease  of  the  hyacinth  the 

writer  has  never  been  able  to  make  satisfactory  thin  sections,  many  of  the  soft  cells 

being  filled  with  bundles  of  very  hard  raphides  which  he  has  not  been  able  to 

dissolve  without  serious  injury  to 

the  tissues.     In  such  cases  thick 

free-hand  sections   are  about  all 

that  can  be  hoped  for. 

Serial  sections  are  cut  on  the 

microtome.     The  one  shown   in 

pi.  13  and  fig.  119  leaves  nothing 

to   be  desired   in  the  way   of  a 

perfect-working  durable  instrument.    The  ribbon-carrier  is  above  the  table  at  the  left. 

The  knife  is  stationary.     The  block  moves  up  and  down,  and  the  razor-carrier 

*Fic.  112. — A  page  from  the  paraffin  record-book.    The  numbers  on  the  slide-boxes  (fig.  103) 
correspond  to  numbers  in  this  book.    Two-thirds  actual  size. 

fFic.  113. — A  mounted  slide  of  serial  sections,  showing  manner  of  labeling. 


ZlVBdL 

B/O.CK  5pot 

DODO 

of    P/U77V. 

X®00 

/rx/.r 

><  xxo 

DOOD 

fVne  cavity 
iijpareijc/ujma 

Fig.  I13.f 


SECTIONS. 


I23 


moves  forward  at  each  stroke  a  distance  governed  by  the  set-screw  of  the  scale 
(y*  J*  to  40  ;").  By  substituting  a  wide  knife-carrier,  sections  several  centimeters  in 
diameter  may  be  cut,  and  by  using  a  slanting  knife,  as  for  celloidin,  very  hard  mate- 
rial may  be  cut.  By  loosening  a  set-screw,  the  razor  as  here  shown  may  be  turned 
a  few  degrees  to  right  or  left,  and  the  paraffin  block  may  also  be  moved  through  a 
considerable  arc  in  any  direction,  it  being  held  securely  in  any  position  by  pressure 
of  a  collar-screw  on  a  ball-and-socket  joint.  On  72  in  plate  13  is  an  apparatus  for 
tracing  the  edges  of  the  paraffin  blocks. 


B 


Fig.  1 14  * 

Collections  of  living  bacteria  are  also  necessary.  Fortunately  many  may  now 
be  obtained,  as  needed,  from  Krai,  in  Prague ;  but,  unfortunately,  they  do  not  always 
correspond  to  their  name.  Others  must  be  kept  on  hand,  and  the  cultures  (of  some 
sorts)  must  be  renewed  at  frequent  intervals.  That  way  which  has  given  the  writer 


*Fic.  114. — A.  Knife  for  serial  sections,  furnished  with  the  Reinhold-'Giltay  microtome.    This 
is  made  by  Joseph  Rodgers  &  Son,  Sheffield,  England.     One-half  actual  size. 

B.  Microtome  knife  made  to  order  by  Charles  Lentz  &  Sons,  Philadelphia,  and  found  useful  in 
cutting  hard  material  with  long  slant  strokes.    One-half  actual  size.    The  broad  wedge-shaped  blade 
of  this  knife  is  shown  in  'ft. 

C.  Knife  obtained  from  J.  R.  Torrey  &  Co.,  Worcester,  Mass.,  and  found  very  useful  for  making 
serial  sections  on  the  microtome.     One-half  actual  size. 

D.  Torrey  razor,  recommended  for  free-hand  sections.    The  very  thin  blade  is  flat  on  one  face 
and  hollow-ground  on  the  other,  as  shown  in  d.     It  is  made  of  the  very  best  steel  and  holds  an  edge 
well.    One-half  actual  size. 

a,  b,  c,  d,  end  views  of  the  cutting  edge  of  knives  shown  in  A,  B,  C,  D.    Actual  size. 


124 


BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 


least  inconvenience  is  by  storage  in  cool  boxes  (refrigerators)  at  temperatures  of  10° 
to  15°  C.  By  tliis  method  some  organisms  can  be  kept  alive  on  agar  a  year  without 
transfer,  and  even  sensitive  organisms  will  generally  live  for  some  months,  especially 


B 

Fig.  115* 

if  planted  in  proper  media.  The  writer  has  never  made  any  attempt  to  prepare  a 
collection  of  dead  bacteria  on  culture  media  to  serve  as  museum  specimens,  but  it 
is  possible  to  do  so,  it  is  said,  with 
considerable  success  by  following  the 
methods  described  by  Hauser  and 
others  (Bibliog.,  L,II). 

DISTILLED  WATER. 

All  laboratories  doing  much  work 
should  have  an  abundance  of  distilled 
water,  and  where  this  is  not  readily 
obtainable  in  sufficient  quantity  and 
of  good  quality,  provision  should  be 
made  for  it  when  the  laboratory  is 
constructed  or  when  the  necessity  for 
it  arises.  In  the  construction  of  such 
a  still  many  things  must  be  kept  in 
mind,  if  it  is  to  work  satisfactorily 
and  yield  water  of  the  desired  purity.  J 


Fig.  116.t 


*Fic.  115.— Cross-section  of  tooth  of  cabbage-leaf  infected  by  Bacterium  campestre.  Plant  No. 
401  sprayed  with  water  containing  an  agar-culture.  Bacterial  occupation  limited  to  points  between 
A  and  B.  At  X  vessels  are  occupied.  At  A  and  B  the  bacteria  lie  in  the  intercellular  spaces  and 
have  not  yet  entered  the  vessels.  For  details  of  A  and  B,  see  figs.  116  and  117.  This  section,  which 
is  one  of  a  series,  was  cut  270  ft  below  the  apex  of  the  leaf-tooth.  A  few  micromillimeters  further 
down  (370  >u)  all  trace  of  the  bacteria  disappears.  In  other  words,  the  bacteria  are  still  confined  to 
the  leaf-tooth,  and  there  is  no  cavity  like  that  shown  in  fig.  76.  When  sprayed  this  leaf  was  extrud- 
ing fluid  from  the  water-pores.  Actual  length  of  section,  slightly  under  I  millimeter.  Slide  331°  3 
Plant  sprayed  December  9,  1904;  slightly  blackened  leaf-tooth  fixed  in  95  per  cent  alcohol  on 
December  17,  1904.  Inked  from  a  photomicrograph. 

fFic.  116. — Cross-section  of  leaf-tooth  of  cabbage  infected  by  Bacterium  catnpestre.  A  detail 
from  fig.  115  at  A.  The  bacteria  have  not  yet  entered  the  vessels. 

tThat  thing  which  has  given  the  writer  most  trouble  was  an  entirely  unexpected  difficulty,  viz, 
a  plague  of  tiny  red  house  ants.  These  got  into  the  reservoir  in  spite  of  all  that  could  be  done  to 
render  it  tight,  and,  of  course,  spoiled  the  water  for  all  delicate  work. 


PLATE  14. 


Apparatus  for  Distilling  Water. 

(I)  Steam  inflow  pips;  (2)  wasts-iteam  pipe  ;  (3)  hydrant- water  inflow  pip:  ;  (4)  hydrant- water  outflow  pipe  (flush)  to 
sewer ;  (5)  galvanized -iron  bailer ;  (6)  water  gage ;  (7)  brass  top,  tinned  on  the  under  side  ;  (6)  copper  catch  basin  ; 
(9)  steam  safety  valve;  (10)  block-tin  steam  pipe  to  condenser;  (11)  block-tin  water  pipe  from  condenser; 
(12)  hydrant- water  pipe  into  condenser  tank;  (13)  hydrant -water  pipe  from  condenser  tank;  (14)  flush  pipe  for 
condenser  tank ;  (1 5)  reservoir,  capacity  80  gallons  ;  (16)  water  gage;  (1  7)  overflow  pipe  from  reservoir ;  (1 8)  block- 
tin  pipe  leading  to  various  rooms ;  (19)  iron  support. 


DISTILLED   WATER. 


125 


The  following  description  and  figure  of  a  distilled-water  apparatus  devised  by 
the  author  for  use  in  the  Laboratory  of  Plant  Pathology,  United  States  Department 
of  Agriculture,  may  be  of  interest,  therefore,  to  some.  The  apparatus  consists  of  a 
galvanized-iron  boiler  similar  to  those  used  in  kitchen  ranges.  It  is  1 8  inches  in 
diameter  and  about  5  feet  high.  The  top  is  sawed  off  and  to  it  is  bolted  a  stout 
iron  ring  with  a  flange,  on  which  rests  a  ^-inch  brass  cover.  In  the  lower  half  of 
this  boiler  is  a  coil  of  52  feet  of  inch  copper  pipe,  the  upper  end  bent  downward 
and  securely  fastened  in  the  bottom  of  the  boiler  to  a  steam  pipe  (i  inch)  connected 
with  a  i  ^4 -inch  steam  pipe  leading  to  the  ordinary  steam  boiler  in  the  engine  room 
in  the  basement;  the  lower  end  connected  with  an  iron  steam  pipe  (i  inch)  leading 
to  a  steam  trap  (Mark  traps  are  said  to  be  the  best).  Around  this  copper  steam 

pipe,  which  is  of  course  tin- 
plated,  stands  the  river  water 
which  is  to  be  converted  into 
steam  by  contact  with  the  hot 
pipe.  This  hydrant  water  is 
kept  always  at  about  the  same 
level  (level  of  fig.  5  in  plate  14), 
by  means  of  a  tinned-copper  ball 
float  (automatic  cut-off)  which 
closes  the  mouth  of  the  inflow 
pipe  when  the  water  rises  be- 
yond a  certain  point.  The  upper 
part  of  the  cylinder  is  a  steam 
chamber  under  very  moderate 
pressure  (o  to  ^  pound,  rarely 
more).  The  excess  of  pressure 
is  dissipated  either  by  escape  of 
steam  through  the  safety  valve 
(9),  which  is  not  weighted,  or 
through  the  coil  of  pipe  in  the 
condenser.  The  steam  passes 
from  a  securely  riveted  tin-lined 

copper  catch  basin  (8)  into  a  ^-inch  block-tin  pipe  (10),  which  is  fastened  to  a 
tubular  projection  from  the  catch  basin  by  means  of  a  collar  screw.  The  tubular 
projection  from  the  top  of  the  catch  basin  is  soldered  in  place  and  also  held  by  a 
flange  inside  the  copper  top,  so  that  it  can  not  be  forced  out  by  any  attainable 
degree  of  steam  pressure.  The  J^-inch  block-tin  pipe  passes  to  the  room  above, 
where  it  is  coiled  for  a  length  of  35  feet  inside  a  tin-lined  copper  tank  resting  on 
the  floor.  The  height  of  the  condensing  tank  is  18  inches  and  its  diameter  is  the 
same.  When  in  operation  this  tank  is  full  of  running  water.  Theoretically,  this 
condensation  tank  is  large  enough,  and  it  is  so  practically  when  the  hydrant  pressure 


*Fic.  117. — Detail  from  fig.  115  at  B,  showing  an  early  stage  of  water-pore  infection  of  cabbage. 
The  bacteria  have  not  yet  entered  the  spiral  vessels.    The  large  dark  bodies  are  nuclei. 


126 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


is  good,  but  when  it  is  feeble  or  when  the  steam  pressure  is  high  the  water  becomes 
too  hot  and  steam  sometimes  escapes  into  the  reservoir.  The  water  therefore  must 
be  hurried  through  the  tank  by  the  use  of  a  steam  pump,  or  else  less  steam  must 
be  allowed  to  enter  the  copper  pipe.  If  the  writer  were  to  build  another  similar 
apparatus  he  would  make  the  condensing  tank  2  feet  higher  and  add  10  feet  to  the 
length  of  the  coil  of  tin  pipe.  The  condensing  tank  is  provided  at  the  bottom 
with  a  i-inch  inflow  pipe  for  the  cold  water  (it  should  be  i^-inch),  and  at  the  top 

with  a  i  "^ -inch  outflow  pipe  (it  should 
be  2-inch),  for  the  exit  of  the  wanned 
water.  There  is  also  a  i-inch  flush 
pipe  at  the  bottom  for  the  occasional 
removal  of  sediment. 

The  size  of  the  outflow  pipe,  which 
must  be  somewhat  larger  than  the  in- 
flow pipe,  prevents  any  possibility  of 
clogging  and  overflow.  All  the  metal 
parts  which  come  into  contact  with  the 
distilled  water  are  tinned  or  nickel- 
plated.  Connected  with  the  lower  end 
of  the  block-tin  coil  (by  tin  solder, 
which  must  not  contain  lead  or  zinc) 
is  a  smaller  (^-inch)  block-tin  pipe 
(i  i ),  which  leads  the  distilled  water  into 
(15)  the  storage  tank  (j^-inch  pipe 
would  be  better,  and  without  any  joint). 
The  reservoir  in  this  case  is  a  white- 
enameled  bath-tub,  on  the  top  of  which 
is  clamped  down  a  cover  of  thin  sheet 
copper  (o'o-inch),  the  inner  face  of 
which  has  been  carefully  tinned.  Plate 
glass  ground  to  fit  would  be  better,  and 
the  tub  itself  is  likely  to  be  discarded 
in  the  near  future,  i.  <?.,  when  some 
more  satisfactory  storage  tank  can  be 
found.  The  problem  of  the  proper 
storage  of  distilled  water  in  quantity  is 
the  hardest  one,  the  solvent  power  of  the 
water  is  so  great.  From  the  bottom  of 
this  bath-tub  several  hundred  feet  of  ^-inch  block-tin  piping  lead  to  various  rooms 
in  the  building.  In  addition  to  the  terminal  faucets  there  is  a  general  cut-off  just 
above  18,  which  is  necessary  in  case  of  an  accident  to  any  faucet  or  part  of  the 
piping.  There  is  also  an  overflow  pipe  (17),  which  does  not  enter  the  sewer,  but 


Fig.  1 18.* 


*Fic.  118. — Early  stage  of  stomatal  infection  in  angular  leaf-spot  of  Rivers  cotton.  Hothouse 
infection  produced  by  spraying  Bacterium  malvacearum  upon  the  surface  of  the  leaves.  For  a  much 
later  stage  see  fig.  80. 


DISTILLED   WATER. 


127 


ends  free  in  the  laboratory  about  i  foot  above  a  deep  sink.  The  sides  and  top  of 
the  boiler,  the  copper  catch  basin,  and  the  -%^-inch  block-tin  pipe  leading  to  the  con- 
denser are  all  coated  with  3  inches  of  best  non-conducting  magnesia  covering. 
The  catch  basin,  designed  to  hold  back  solid  particles  carried  up  with  the  steam, 
is  9  by  12  inches  and  is  made  of  ^ -inch  copper,  securely  riveted  and  soldered  with 
tin  solder.  It  is  bolted  down  to  the  flat  brass  top  and  a  steam-tight  connection 


Fig.  1 19.* 

is  secured  by  means  of  a  red  rubber  gasket.  The  heavy  brass  top  (7)  is  tinned  on 
the  inner  surface  and  is  bolted  securely  to  the  iron  flange  on  the  top  of  the  boiler  by 
means  of  1 8  screw-bolts.  The  junction  is  made  steam-tight  by  means  of  a  corrugated 

*FiG.  119. — The  Reinhold-Giltay  microtome  arranged  for  cutting  celloidin  or  very  hard  paraf- 
fin sections.  The  machine  is  very  solidly  and  accurately  constructed  out  of  the  best  materials, 
and,  in  addition,  provision  is  made  by  means  of  set-screws  for  compensating  the  wear  due  to  long 
use.  The  device  governing  the  thickness  of  the  sections  is  especially  ingenious.  This  particular 
machine  has  been  in  constant  use  by  various  persons  for  over  four  years,  and  nothing  has  been  paid 
out  for  repairs.  With  good  use  it  ought  to  last  a  lifetime.  About  one-fifth  actual  size. 


128  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

tinned-copper  gasket.  The  steam  which  runs  the  apparatus  is  brought  to  the  lab- 
oratory floor  through  a  i^-inch  pipe,  in  which  (in  the  engine  room)  there  is  a 
steam  gage  registering  up  to  150  pounds,  and  a  reducing  valve  set  at  55  pounds. 
This  very  considerably  lessens  the  steam  pressure  in  the  copper  coil,  moderates  the 
violence  of  the  ebullition,  and  makes  the  apparatus  perfectly  safe.  The  hydrant- 
water  outflow  pipe  (flush)  to  the  sewer,  for  occasionally  washing  out  accumulated  mud 
(4)  passes  from  the  bottom  of  the  boiler  immediately  above  fig.  19.  Gate-valves 
are  used.  All  brass  and  copper  parts  in  contact  with  the  steam  are  tinned;  all 
metal  parts  in  contact  with  the  distilled  water  are  tin,  tinned,  or  nickel-plated. 

With  60  pounds  steam  pressure  in  the  engine-room  boiler,  40  pounds  pressure 
at  the  reducing  valve,  35  pounds  pressure  in  the  pipe  at  the  laboratory  floor  near 
where  it  enters  the  still,  and  one-half  pound  pressure  or  less  in  the  steam  chamber 
above  the  coil  of  copper  pipe,  the  capacity  of  this  still  is  60  liters  ( 1 6  gallons)  per  hour. 

The  apparatus  must  be  built  very  substantially  in  all  parts,  so  as  to  withstand 
at  least  twice  as  much  steam  pressure  as  any  part  of  it  will  be  subjected  to,  e.g.-,  160 
pounds  in  the  iron  pipes  and  in  the  copper  coil  and  its  attachments,  and  at  least  20 
pounds  in  the  catch  basin,  and  other  parts  subject  to  steam  generated  in  the  still. 
A  steam  gage,  in  addition  to  the  one  in  the  engine-room,  shows  the  pressure  in 
the  coils,  and  another  the  pressure  in  the  steam  chamber  above  the  coils.  They  are 
not  shown  in  the  plate,  as  they  were  put  on  after  that  was  made.  The  former  is 
attached  to  the  steam  supply  pipe  near  the  floor,  and  the  latter  to  an  arm  of  the 
safety-valve  pipe.  The  boiler  should  be  taken  down  and  the  parts  retinned  once  a 
a  year,  or  at  least  once  in  two  years. 

If  a  much  greater  quantity  of  water  is  needed  the  block-tin  condensation  coil 
should  be  lengthened  to  60  feet,  the  diameter  of  the  inflow  pipe  of  the  condenser 
should  be  increased  to  2  inches,  and  the  outflow  pipe  to  2  %  inches,  and  the  cubic 
contents  of  the  condenser  tank  should  be  quadrupled.  The  capacity  of  the  bath-tub 
(or  other  receptacle),  for  a  large  laboratory  should  be  at  least  500  liters,  and  might 
well  be  i, ooo  liters. 

The  above  apparatus  has  been  in  use  for  two  years.  It  works  very  smoothly  and 
satisfactorily  when  the  proper  amount  of  steam  is  let  into  the  coil  of  copper  pipe, 
which  ordinarily  should  not  be  nearly  the  whole  amount  available.  The  inflow  of 
steam  is  governed  by  the  valve  a  few  inches  below  fig.  i  in  plate  14.  When 
too  much  steam  enters  the  coil,  the  pressure  in  the  steam  chamber  above  it  rises  to 
five  pounds  or  more,  hot  water  is  forced  back  through  the  feed  pipe  (3)  into  the 
neighboring  pipe  which  furnishes  cold  water  to  the  condenser  (12),  and  steam  in- 
stead of  distilled  water  is  furnished  to  the  water  tank.  This  is  at  once  obviated  by 
cutting  off  part  of  the  steam  inflow  and  moderating  the  force  of  the  boiling.  It 
might  also  be  obviated  by  reducing  the  length  of  the  arm  of  the  safety  valve  (9), 
which  in  any  event  should  not  be  weighted. 

Sufficient  water  for  small  quantities  of  culture-media  and  pure  enough  for  most 
purposes  may  be  obtained  from  the  simple  glass  still  shown  in  fig.  82  by  one  dis- 
tillation. Water  of  a  high  degree  of  purity  may  be  obtained  by  two  distillations, 
adding  0.5  gram  to  i  gram  of  potassium  permanganate  per  liter  of  water  before  the 


NON-SOLUBLE   GLASS. 


129 


first  distillation,  and  5  grams  of  c.  p.  sulphuric  acid  per  liter  before  the  second 
distillation.  The  flasks  in  which  such  water  is  collected  or  stored  should  be  of 
resistant  (non-soluble)  glass  and  absolutely  clean  to  begin  with.  With  use  such 
flasks  or  bottles  become  more  valuable  and  should  not  be  employed  for  other  purposes. 

The  solubility  of  glassware  is  best  tested  by  determining  from  time  to  time 
the  degree  of  electrical  conductivity  of  pure  water  stored  in  it.  The  specific  resist- 
ance of  pure  water  stored  for  a  week  in  such  tubes,  flasks,  or  bottles  should  not  fall 
below  250,000  ohms.  The  specific  electrical  resistance  is  determined  upon  i  cubic 
centimeter  of  water  exposed  between  electrodes  having  an  area  of  i  square  centi- 
meter, and  is  read  by  means  of  a  special  Wheatstone  bridge.  Distilled  water 
redistilled  with  chromic-acid  cleaning  mixture,  and  afterwards  with  alkaline  potas- 
sium permanganate  (method  used  by  the  Physical  Laboratory  in  the  Bureau  of 
Soils)  gives  a  resistance  of  700,000  ohms. 

The  following  determinations  made  by  the  Physical  Laboratory  of  the  Bureau 
of  Soils  show  the  diverse  behavior  of  two  lots  of  clean  test-tubes  recently  purchased 
as  non-soluble  glass  by  the  Laboratory  of  Plant  Pathology. 


Kind  of  tube. 

Time  of  exposure, 
iu  days. 

Specific  resistance, 
iu  ohms. 

Resistant  test-tubes,  (R)  from  Greiner 

Do     2d  test  

Tubes  received  from  the  School  Sup- 
olv  Co  .  . 

Do.  ,  zd  test  

The  twice-distilled  water  used  was  taken  from  a  Jena  flask  and  its  initial 
specific  resistance  was  240,000  ohms. 

MICROSCOPES. 

Microscopes  of  a  much  better  grade  are  required  for  bacteriological  investigations 
than  for  ordinary  histological  work.  The  writer  has  for  many  years  employed  those 
made  by  Carl  Zeiss,  of  Jena,  as,  on  the  whole,  most  serviceable.  Good  microscopes 
are  also  made  by  E.  Leitz,  of  Wetzlar,  and  recently  by  the  Spencer  Lens  Company, 
of  Buffalo,  N.  Y.  The  Zeiss  stand  shown  in  plate  15  does  very  well  for  all  ordinary 
work,  but  is  not  well  adapted  for  the  making  of  photomicrographs  or  for  recording 
the  exact  location  of  particular  spots  in  the  section.  The  latter  difficulty  may, 
however,  be  overcome  by  means  of  a  removable  slide-carrier  attached  to  the  stage. 
The  stand  may  also  be  used  with  the  small  upright  photomicrographic  outfit  shown 
in  fig.  24  when  the  lens  does  not  require  a  microscope  having  a  wide  tube  This 
microscope  has  a  half-mechanical  stage,  an  excellent  fine  adjustment,  and  good 
substage  apparatus.  It  is  thoroughly  well  made  and  very  durable.  One  in  the 
writer's  laboratory  has  been  in  use  for  twelve  years.  The  lacquer  has  disappeared 
in  places  and  it  is  no  longer  attractive  to  look  at,  but  it  has  required  no  serious 
repairs  during  this  time  and  is  still  serviceable. 

For  photomicrographic  work  and  also  for  recording  the  exact  location  of  desir- 
able fields  in  a  section,  the  writer  uses  the  large  photomicrographic  stand  shown  in 
plate  1 6.  This  is  provided  with  a  specially  wide  barrel,  a  fine  adjustment  of  very 


PLATE  IS. 


Zeiss  microscope  stand  II". 

This  form  of  microscope  and  that  represented  on  plate  16  are  the  two  patterns  used  princi- 
pally in  the  Laboratory  o(  Plant  Pathology,  U.  S.  Department  of  Agriculture.  The 
objectives  are  apochromatic.  and  have  proved  very  serviceable.  In  carrying  do  not  grasp  by 
any  part  above  the  level  of  the  stage,  as  this  brings  an  undue  weight  upon  the  fine  adjust- 
ment. Seize  by  the  base. 


PLATE  16  . 


Zeiss  photomicrographic  stand  Ic. 

The  barrel  "T"  is  of  greater  diameter  than  in  stand  lla.  The  fine  adjustment  is  at  "  W"  and  no 
weight  rests  on  it  in  lifting  the  instrument  by  the  handle  "  H."  The  set  screw  "  K  "  locks  the  upper 
part  of  the  instrument  at  any  angle.  The  objective  is  set  in  place  by  means  of  a  very  convenient  slide 
carrier.  The  fine  adjustment  screw  has  an  extremely  slow  movement  ;  and  the  vernier  screws  are  on 
the  same  axis  (a  great  convenience).  The  stage  rotates  and  may  be  locked  at  the  desired  place  by 
means  of  a  set  screw.  For  the  subctage  arrangement  see  figure  120- 


130 


BACTERIA    IN    RELATION   TO    PLANT   DISEASES. 


slow  movement,  a  swing-out  condenser  (fig.  120),  two  substage  iris  diaphragms,  and 
various  other  conveniences.  For  example,  the  screw-heads,  determining  the  cross 
and  sidewise  movement  of  the  section,  are  on  the  same  axis  and  may  be  reached  and 
moved  without  changing  the  position  of  one's  arm. 

The  apochromatic  objectives  are  the  only  ones  recommended  for  bacteriological 
work.  They  cost  more  than  achromatic  objectives,  but  expense  is  a  minor  con- 
sideration. In  hot,  moist  climates  the  older  apochromatic  objectives  of  Zeiss  fre- 
quently became  clouded,  but  those  made  in  recent  years  have  given  the  writer  no 
trouble  in  the  latitude  of  Washington.  They  yield  sharp  images  even  with  high 
eye-pieces.  Of  course,  compensating  oculars  must  be  used  with  the  apochromatic 
objectives.  It  is  de-  ™) 

sirable  to  have  the 
whole  series  of  ob- 
jectives and  eye- 
pieces, but  if  one  is 
limited  for  means, 
very  good  work  can 
be  done  with  two 
objectives  and  three 
oculars,  viz,  object- 
ives 1 6  mm.  and  3 


mm.  1.40  n.  a., 


and 


compensating  oculars 
4,  6,  and  12. 

The  newer  forms  of  the  Abbe 
camera  furnished  by  Zeiss  (fig.  121) 
leave  little  to  be  desired  in  the  way 
of  a  drawing  camera. 

PHOTOGRAPHY   AND   PHOTO- 
MICROGRAPHY. 

For  permanent  records  nothing 
equals  photography.  It  constitutes, 
therefore,  a  very  important  special  Flg-  l20'* 

part  of  laboratory  work,  and  every  student  of  pathology  should  make  a  knowledge 
of  this  subject  part  of  his  education.  Some  of  the  following  suggestions  will  be 
useful  to  beginners. 

The  Zeiss  Double-Protar  lenses,  series  Vila,  are  the  best  all  round  photographic 
lenses  made  by  that  firm,  and  are  excelled  by  none  made  by  any  firm.  The  back 
or  front  lens  is  usually  as  good  as  the  combination.  Excellent  photographic  lenses 
are  also  made  by  Voigtlaender  and  by  Goerz.  Zeiss  photographic  lenses  may  be 

*Fic.  120. — Swing-out  condenser  and  other  substage  arrangements  on  Zeiss  photomicrographic 
stand,  No.  ic.  There  is  an  iris  diaphragm  in  D,  and  a  second  one  in  S,  which  is  for  use  when  the 
condenser  is  thrown  out  as  shown  in  this  figure.  D  swings  under  when  C  is  thrown  into  place. 
W  racks  the  entire  substage  up  or  down. 


PHOTOGRAPHY  AND    PHOTOMICROGRAPHY. 

obtained  from  Bausch  &  Lomb,  who  are  under  contract  to  manufacture  them 
according  to  the  Zeiss  fonnulse.  In  buying  a  photographic  outfit  it  is  economy  to 
get  one  of  the  high-priced  lenses.  It  is  frequently  stated,  by  those  who  do  not 
know,  that  "just  as  good  results"  can  be  obtained  with  cheap  lenses,  but  one  may 
easily  satisfy  himself  that  such  is  not  the  case  by  photographing  buildings  on  a 


Fig.  121.- 

street  or  any  object  having  many  vertical  parallel  lines  and  other  lines  crossing  at 
right  angles.  The  pictures  made  by  the  cheap  lenses  generally  show  serious  distor- 
tions. In  buying  a  lens  one  should  know  in  advance  exactly  what  he  wishes  to  do 
with  it,  otherwise  he  may  be  greatly  disappointed.  If  he  wishes  to  photograph  only 


*Fic.  121. — Newer  form  of  Zeiss-Abbe  drawing  camera.  The  camera  is  clamped  at  K  by  means 
of  S.  The  prism  within  R  is  centered  over  the  eye-piece  by  screw  movements  of  L  and  Z.  When 
not  in  use  the  prism  is  swung  to  the  right,  as  indicated  by  the  dotted  lines.  The  mirror  A  throws 
down  the  prismatic  image  to  the  drawing  paper.  The  amount  of  light  is  governed  by  the  substage 
iris-diaphragm  and  by  rotating  B  and  R,  which  contain  smoky  glasses  of  graded  densities.  P  is 
an  extra  prism.  The  image  on  the  paper  will  also  be  clearer  if  it  is  placed  in  shadow  by  means  of 
a  screen  of  some  sort 


132 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


flat  surfaces  he  will  select  a  lens  with  no  great  penetration,  but  with  a  very  clear  field, 
sharp  to  the  edges,  i.  e.,  a  Planar  or  some  similar  lens.  If  he  needs  a  lens  with 
very  little  depth  of  focus  (but  more  than  the  Planar)  and  one  allowing  dark  objects 
to  be  photographed  in  a  very  short  time,  e.g.,  luminous  bacteria  by  their  own  light, 
he  will  select  a  Zeiss  Unar  or  its  equivalent,  i.  e.,  an  extremely  rapid  lens.  If  he 
desires  in  one  picture  as  much  as  possible  of  a  landscape,  e.g.,  a  large  tree  or  an 
interior,  he  will  select  an  extremely  wide-angle  lens  rather  than  one  distinguished 
for  its  rapidity  or  for  the  perfection  of  its  definition,  e.  g.,  a  Zeiss  Protar,  series  V. 
The  Double-Protar,  series  Vila,  combines  as  wide  an  angle,  as  flat  a  field,  as  great 
rapidity,  and  as  sharp  a  definition  as  it  is  possible,  apparently,  to  obtain  in  a  lens 
and  at  the  same  time  have  great  depth  of  focus.  These  lenses  may  also  be  unscrewed 
and  each  half  used  separately,  if  one  wishes  some  portion  of  a  picture  more  highly 
magnified.  They  are  furnished  with  front  and  back  lenses  of  equal  or  unequal  focal 
distance,  as  may  be  desired. 

In  using  Planars  and  all  lenses  which  magnify,  it  is  necessary  to  secure  a  very 
exact  focus  with  the  stop  wide  open,  for,  unlike  lenses  which  give  pictures  less  than 


Fig.  122* 

actual  size,  only  a  very  little  increased  depth  of  focus  can  be  obtained  by  stopping 
down.  With  many  objects — e.  g.,  the  surface  of  a  leaf,  or  of  bacterial  colonies — 
there  is  considerable  difficulty  in  deciding  which  is  the  proper  focus  when  a  Planar 
is  used,  what  seemed  like  a  good  focus  often  yielding  a  poor  negative.  On  this 
account  the  writer  is  in  the  habit  of  focusing  on  a  fragment  of  very  fine,  sharp 
print  laid  on  the  surface  of  the  leaf  or  of  the  agar-plate  near  the  colonies  to  be 
photographed.  A  lens  magnifying  6  times  is  used  in  judging  of  the  image  on  the 
ground  glass,  and  when  the  best  possible  focus  has  been  secured,  the  paper  is  removed, 
the  lens  is  stopped  down  two-thirds,  and  the  photograph  is  made.  In  case  of  white 
colonies  the  best  results  are  obtained  by  resting  the  Petri  dish  on  a  piece  of  black 
paper  while  the  photograph  is  being  made.  The  exposure  is  shortened  by  illumi- 
nating the  surface  of  the  object  with  a  bright  beam  from  a  mirror.  The  apparatus 

*Fic.  122. — Zeiss  Planar  lenses,  series  la,  Nos.  i  to  5.  Nos.  i,  2,  and  3  may  be  attached  to  the 
funnel-shaped  carrier  shown  in  the  figure.  This  screws  into  the  top  of  the  microscope  barrel  in 
place  of  the  eye-piece  tube.  The  one  attached  is  No.  3.  The  condensing  lenses  necessary  for  these 
Planars  are  also  shown  in  this  figure,  at  right  and  left. 


PHOTOGRAPHY   AND    PHOTOMICROGRAPHY. 


133 


shown  in  fig.  24  may  be  used  for  this  purpose.  To  avoid  shadows  the  mirror  should 
be  held  some  distance  above  the  object  when  the  surface  is  not  even.  The  first  five  of 
the  Zeiss  series  of  Planars  are  all  that  are  usually  required.  No.  i  gives  the  highest 


Fig.  123.* 


*Fic.  123. — Simple  apparatus  for  holding  the  camera  in  place  when  one  wishes  to  photograph 
down.  The  camera  here  shown  is  a  Rochester  Optical  Company,  reversible  back  5  by  8,  fitted  with 
a  Bausch  &  Lomb  rapid  universal  lens,  and  has  been  used  very  often  by  the  writer  for  natural-size 
work  and  for  lantern  slides. 


134 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


magnification ;  No.  5,  the  largest  field  ; 
No.  3  will  give  a  sharp  image  of  a  flat 
obj  ect  a  centimeter  in  d  iameter.  Special 
condensing  lenses  are  required.  These 
fit  into  the  snbstage  in  place  of  the 
Abbe  condenser.  One  condenser  serves 
for  Nos.  1,2,  and  3,  and  another  for  Nos. 
4  and  5  (fig.  122). 

In  photographing  poured-plate  colo- 
nies natural  size,  there  are  several  ways. 
It  may  be  done  by  reflected  light,  as 
shown  in  fig.  123,  in  which  case  the 
colonies  sometimes  cast  deep  shadows. 
Such  shadows  may  be  avoided  by 
mounting  the  camera  as  shown  in  fig. 
1 24  and  gently  twirling  it  during  the 
exposure.  The  Petri  dish  may  also  be 
photographed  by  transmitted  light  ex- 
actly as  if  it  were  a  negative  for  a 
lantern  slide.  The  Petri  dish  is  then 
held  in  place  in  the  darkened  window 
or  in  front  of  the  camera  box  by  crowd- 
ing it  into  a  hole  cut  in  a  square  of 
thick  leather,  paper,  or  sheet-rubber 
(^  inch),  which  is  then  fastened  over 
the  kit  or  framework  by  eight  thumb- 
tacks, or,  better,  it  may  be  held  in  place 
by  two  stout  rubber  bands,  as  shown  in 
the  photographs  (plate  17  and  fig.  125). 
With  stop  32  u.  s.  and  Seed's  2/-X 
plates  the  right  exposure  in  Washing- 
ton is  usually  somewhere  between 
^  second  and  \  second  in  sunny  weather 
and  3  to  5  seconds  in  cloudy  weather, 
using  a  Voigtlaender  collinear  lens, 
series  III,  No.  6,  and  south  light. 

Atkinson  gets  very  good  results  by 


Fig.  124.* 


*Fio.  124.— Modified  Collins-Brown  camera  swung  from  the  ceiling  and  set  to  magnify  about 
X  1 34-  The  four  suspending  strings,  which  are  of  very  strong  fish-line,  end  in  an  S-shaped  hook, 
the  upper  end  of  which  hooks  over  a  ring  attached  to  a  stout  cord  pendant  from  the  ceiling.  The 
length  of  bellows  in  this  camera  as  modified  by  the  writer  is  25  inches.  The  lens  used  with  it  is  a 
Zeiss  Double-Protar,  Series  Vila.  No.  13,  made  by  Bausch  &  Lomb,  Rochester,  N.  Y.  This  is  the 
type  of  lens  known  also  as  the  Zeiss  Convertible  Double  Anastigmatic.  This  lens  has  a  focal  dis- 
tance of  pJ4  inches,  or,  when  only  the  front  or  back  half  is  used,  16^2  inches  (16  according  to  Zeiss 
catalogue).  It  is  provided  with  a  Bausch  &  Lomb  No.  2  Volute  shutter.  A  cork  support  was 
placed  under  the  object  carrier  to  steady  the  apparatus  while  it  was  being  photographed,  but  in 
actual  use  the  camera  swings  free,  and  if  one  desires  to  avoid  shadows  the  apparatus  is  given  a 
gentle  twirl  just  as  the  exposure  begins.  The  object  carrier  is  easily  removed,  and  is  held  in  place 
at  any  level  by  two  set-screws. 


PLATE   17. 


Enlarging  and  reducing  camera,  showing  method  of  mounting  the  apparatus. 

On  the  table  at  the  left  is  a  Petri-dish  poured  plate  held  in  place  by  two  rubber  bands  and  ready  for  photographing.    On  the  table  at  the  right  is  a  specia 
camera-back  used  in  making  lantern  slides.     This  allows  the  ground  glass  to  be  railed  or  lowered,  pushed  to  right  or  left,  or  rotated  at  will 


PHOTOGRAPHY   AND    PHOTOMICROGRAPHY. 


135 


placing  a  circular  black  disc  centrally  some  distance  behind  the  plate  to  be  plioto- 
graphed,  using  for  illumination  the  diffused  light  which  conies  in  around  this  disc. 
The  result  is  a  very  sharp  contrast,  i.  e.,  white  colonies  on  a  black  background 
(Bull.  Torrey  Bot.  Club,  1893,  Vol.  XX,  p.  357). 

In  photographing  test-tube  cultures  the  chief  trouble  is  the  great  number  of 
confusing  high-lights  due  to  the  curved  surface  of  the  glass.  From  an  artistic 
standpoint  these  are  to  be  desired,  but  inasmuch  as  they  are  sometimes  liable  to  be 
mistaken  for  bacterial  growths  the  naturalist  desires  to  eliminate  them.  This  may 


Fig.  125* 

be  done  in  several  ways.  One  of  the  best  ways  is  to  photograph  the  tubes  through 
a  thin  sheet  of  distilled  water.  For  this  purpose  jars  of  clear  white  glass  are 
necessary.  These  should  be  about  5^  wide  X  5f  deep  X  i  ^  inches  thick  (inside 
measure),  with  parallel  walls  and  a  flat  bottom.  Such  jars  may  be  obtained  of  Emil 
Greiner.  Only  those  without  flaws  or  wavy  lines  should  be  accepted.  Better  jars 
with  perfectly  parallel  flint-glass  walls  may  be  had  from  Carl  Zeiss.  Good  results 


*Fic.  125. — Showing  method  of  holding  Petri-dish  poured  plates  for  photographing  by  trans- 
mitted light  with  camera  shown  in  plate  17.  The  dish  Is  held  in  place  by  two  stretched  rubber 
bands,  exactly  as  if  it  were  a  negative  to  be  used  for  making  lantern  slides.  For  manner  of  sus- 
pension, »'.  e.,  relation  to  the  camera,  see  plate  17.  The  organism  is  a  48-hour  agar  culture  of  van 
Hall's  Ps.  syringae  II,  grown  at  24°  to  27°  C. 


136  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

may  also  be  obtained  by  photographing  the  tubes  against  a  north  light  inside  a 
box  with  blackened  walls.  The  box  may  be  8  X  8  X  12  inches,  open  at  each  end, 
and  painted  inside  with  a  mixture  of  lamp-black  and  turpentine.  One  open  end  is 
pointed  to  the  window,  the  other  to  the  lens  of  the  camera.  In  the  middle  of  the 
box,  crosswise,  on  top,  a  row  of  ^-inch  holes  (i^  inches  apart)  is  bored,  and  the 
tubes  to  be  photographed  are  thrust  through  these.  If  images  of  outside  objects 
appear  on  the  ground  glass,  they  may  be  cut  out  by  pasting  white  tissue-paper  on 
the  window-glass  (this  should  be  glued  only  at  the  corners). 

The  kind  of  plate  used  depends  upon  the  object.  If  the  contrasts  are  very  great — 
e.  g.,  a  waterfall  or  bright  rock  surrounded  by  vegetation,  or  an  interior  with  the 
camera  pointed  toward  windows — a  double-coated  non-halation  plate  should  be  used 
(Hammer's  Aurora  plates  are  very  good ;  plate  6  was  made  with  such  a  plate).  In  the 
absence  of  such  plates  most  of  the  halation  may  be  avoided  by  squeegeeing  to  the 
back  of  the  dry  plate,  before  loading,  a  black  paper  soaked  in  glycerin  (plate  14  was 
made  in  this  way).  The  dry  plate  should  be  placed  face  down  on  dry  blotting  paper 
during  this  process,  and,  of  course,  the  glycerin-soaked  paper  must  be  cut  in  advance 
to  fit  the  dry  plate.  Much  may  be  done  during  development  to  avoid  violent  con- 
trast if  one  knows  how,  the  quantity  of  pyrogallol  or  ortol  being  greatly  reduced  and 
the  development  prolonged.  This  gives  a  thin  negative  full  of  detail. 

For  many  purposes  isochromatic  plates  are  invaluable ;  for  other  purposes  ordi- 
nary plates  will  give  better  results.  Which  kind  is  best  adapted  to  a  particular 
subject  will  depend  on  what  is  wanted.  In  general,  for  this  sort  of  work,  the  full 
contrast  of  the  original  is  desired,  and  the  kind  of  plate  which  will  give  it  is  best. 
Even  some  exaggeration  of  the  contrasts  of  the  original  is  not  objectionable  in  many 
cases  if  the  prints  are  to  be  used  for  half-tone  reproductions,  since  contrast  is  often 
reduced  in  the  half-tone  process,  and  there  must  be  exaggeration  in  the  photograph 
if  the  half-tone  picture  is  to  correctly  represent  the  object.  An  example  or  two  will 
help  the  beginner  to  judge.  If  we  have  black  spots  on  a  green  background  and 
use  a  rapid  non-isochromatic  plate,  we  will  get  the  result  shown  in  fig.  1 26,  which 
may  be  contrasted  with  fig.  127,  made  from  the  same  leaf,  but  on  an  isochromatic 
plate.  For  such  cases  the  isochromatic  plate  is  of  course  the  one  to  be  selected.  In 
the  same  way,  if  one  desired  to  reproduce  the  variegations  of  a  pansy  with  the  full 
value  of  each  color,  he  would  use  an  isochromatic  plate.  On  the  contrary,  white 
spots  or  stripes  on  a  green  leaf,  or  yellow  colonies  on  an  agar  or  gelatin  plate,  or  red 
spots  on  a  white  ground,  will  stand  out  better  if  the  photograph  is  made  on  a  Seed's 
2J-X  non-isochromatic  plate  or  its  equivalent.  Red  spots  on  a  green  background 
require  an  isochromatic  plate.  Black  spots  on  a  yellow  or  orange  ground  usually 
require  for  good  contrasts  an  isochromatic  plate.  Some  yellows,  however,  take  pale, 
while  others  take  dark. 

In  making  photomicrographs  little  trouble  is  experienced  with  low  powers,  but 
there  is  considerable  difficulty  in  making  good  negatives  of  bacteria  in  tissues,  using 
high  powers.  A  few  hints  may  be  of  service.  With  upright  stands  and  certain 
objectives  the  beginner  frequently  has  difficulty  in  securing  a  uniformly  lighted 
field.  This  trouble  may  be  obviated  by  throwing  the  light  from  the  mirror  not 


PHOTOGRAPHY   AND    PHOTOMICROGRAPHY.  137 

directly  on  the  substage  mirror  of  the  microscope,  but  on  a  sheet  of  ground  glass 
(it  may  be  the  focusing  plate  of  the  camera)  placed  in  front  of  the  mirror  of  the 
microscope.  The  coarse  adjustment  of  the  microscope  should  not  work  too  easily, 
or  else  the  mere  weight  of  the  microscope  tube  may  throw  out  the  focus  after  it  has 
been  secured  and  before  the  picture  can  be  taken.  The  connection  between  camera 
and  microscope  must  be  light-tight.  In  absence  of  a  proper  device  (foot  of  stand 
in  fig.  24),  light  may  be  cut  out  by  several  folds  of  black  velvet  pinned  close.  The 
stage  of  the  microscope  should  also  be  protected  from  bright  reflected  light  when 
photographing  by  transmitted  light.  If  there  is  a  rigid  connection  between  the 
camera  and  the  top  of  the  microscope,  or  if  the  latter  rests  on  the  base  of  the  former, 
the  focus  is  apt  to  be  injured  by  slight  jars  incident  to  putting  in  the  plate-holder 
or  drawing  the  slide.  For  this  reason  it  is  better  to  have  them  separate,  and  the 
carrier  and  draw-slide  should  be  scraped,  sandpapered  or  filed,  and  waxed,  soaped, 
or  vaselined,  so  as  to  work  very  smoothly.  An  entire  day  spent  in  accomplishing 
this  end  should  not  be  counted  as  wasted  time. 

With  large  horizontal  cameras  (plate  5)  the  work-table  and  the  bellows-table 
must  be  leveled  up  accurately  with  reference  to  each  other,  sidewise  as  well  as 
vertically,  and  then  must  be  bolted  to  the  floor.  The  order  of  apparatus  beginning 
at  the  window  is :  mirror,  condensing  lens,  alum-cell,  light-filter  (Zettuow's  fluid),* 
microscope,  automatic  shutter,  front  board  of  the  camera,  large  black  diaphragm  in 
middle  part  of  bellows,  ground  glass  of  the  camera.  The  newer  styles  have  a  screw- 
device  for  elevating  or  lowering  the  camera  and  another  for  elevating  or  lowering 
the  microscope,  or  the  optical  bench.  Dr.  Novy  has  added  to  his  Zeiss  table  a  very 
convenient  device  by  means  of  which  the  services  of  an  assistant  are  dispensed  with, 
one  person  behind  the  ground  glass  of  the  camera  being  able  from  this  position  to 
move  the  slide  in  any  direction  desired.  The  cost  of  the  attachment  is  about  $15. 

Beginning  with  the  center  of  the  mirror  at  the  far  end  of  the  work-table  or 
beyond  it,  and  ending  with  the  center  of  the  ground  glass  at  the  back  of  the  camera, 
all  parts  of  the  apparatus  must  be  centered  accurately,  i.  e.,  the  light  reflected  from 
the  center  of  the  mirror  must  pass  in  a  straight  line  through  the  center  of  the  con- 
densing lens,  Abbe  condenser,  objective,  and  eye-piece  to  the  center  of  the  ground 
glass  at  the  back  of  the  camera,  otherwise  a  first-class  negative  will  not  be  obtained. 
The  Abbe  condenser  must  also  be  at  the  right  distance  from  the  stage  of  the  micro- 
scope ;  the  image  will  then  be  on  the  center  of  the  ground  glass,  circular,  uniformly 
lighted,  and  free  from  distortion  and  color  fringes,  if  the  optical  parts  are  in  proper 
working  order.  The  distance  of  the  Abbe  condenser  varies,  of  course,  with  the 
objective.  The  Planar  lenses  require  special  substage  condensers,  such  as  those 
shown  in  fig.  122.  When  the  centering  is  perfect  all  the  rest  is  easy,  or  becomes 
easy  with  a  little  experience.  If  sunlight  is  used,  an  automatic  shutter  should  be 
placed  on  the  end  of  the  camera  next  the  microscope,  so  that  accurately-timed  short 
exposures  may  be  made.  The  sun's  rays  should  pass  through  several  inches  of  fluid 

*This  is  a  mixture  of  copper  nitrate  and  chromic  acid  in  distilled  water.  It  lets  through  only 
the  greenish-yellow  rays.  This  fluid  acts  on  the  cement  of  the  flint-glass  container,  and  should 
not,  therefore,  be  allowed  to  stand  in  it  longer  than  necessary.  The  latter  should  then  be  washed 
in  pure  water  and  properly  drained. 


i38 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


before  they  enter  the  objective ;  otherwise,  if  the  focus  of  the  condensing  lens  should 
accidentally  coincide  with  the  balsam  mount  of  the  lenses  for  a  few  minutes,  it  may 
be  softened  and  the  objective  ruined.  Pure  water  is  as  good  for  this  purpose  as  alum 
water,  which  was  formerly  much  recommended.  It  removes  more  than  50  per  cent 
of  the  heat  rays. 

The  writer  uses  a  Zeiss  3-inch  mirror  with  micrometer-screws  for  throwing  the 
sun's  rays.     This  serves  quite  as  well  as  the  more  expensive  heliostat,  if  one  can 


Fig.  126.* 

work  quickly.  The  order  ot  procedure  is  to  obtain  the  proper  focus  and  see  that  it 
"  holds  ; "  the  plate  holder  is  then  introduced  and  opened,  and  consequently  the  bel- 
lows must  be  light-tight ;  last  of  all,  the  sunlight  is  accurately  re-centered  and  the 
shutter  snapped.  The  photomicrograph  should  be  made  with  light  from  the  central 

*Fic.  126. — Fragment  of  a  green  leaf  bearing  black  spots.  Enlarged  &A  times  with  a  Zeiss 
Planar  lens  and  photographed  on  a  Seed's  27-X  plate.  Introduced  for  comparison  with  fig.  127. 
Notice  that  although  stopped  down  considerably,  part  of  the  leaf  is  out  of  focus. 


PHOTOMICROGRAPHY. 


139 


portion  of  a  considerable  image  of  light.  My  custom  is  to  nearly  close  the  iris 
diaphragm  below  the  Abbe  condenser  and  throw  with  the  condensing  lens  a  small 
circle  of  light  into  the  center  of  this  diaphragm  ;  the  condensing  lens  is  then  slid 
along  the  track  about  12  or  15  inches  nearer;  the  iris  diaphragm  is  then  opened 
wide  and  the  exposure  made  at  once  by  squeezing  the  bulb  of  the  shutter. 

I  now  always  use  apochromatic  lenses  and  never  make  negatives  without  an 
eye-piece.     I  have  used  Zeiss  projection  oculars,  but  now  use  in  preference  a  Zeiss 


Fig.  127.* 

No.  4  compensating  ocular,  or  Spencer  No.  3,  which  is  kept  solely  for  this  purpose 
(so  as  to  be  always  clean).  It  is  of  the  utmost  importance  that  mirror,  walls  of 
light-filter,  alum-cell,  and  surfaces  of  condenser,  slide,  objective,  and  ocular  be  abso- 
lutely free  from  dirt,  grease,  and  dust  particles,  even  the  smallest,  if  a  good  negative 

*Fic.  127. — Bacterial  leaf-spot  of  the  larkspur  (Delphinium).  Same  as  fig.  126,  but  photo- 
graphed on  Cramer's  isochromatic  slow  plate.  In  this  photograph  the  black  spots  on  a  green  back- 
ground come  out  distinctly;  in  fig.  126  they  do  not 


140  BACTERIA    IN    RELATION   TO    PLANT   DISEASES. 

is  desired.  When  using  oil-iminersion  objectives  see  that  there  are  no  air-bubbles 
or  particles  of  dirt  in  the  cedar  oil.  The  image  on  the  ground  glass  should  be 
observed  the  last  thing  before  introducing  the  plate-holder,  to  see  that  it  is  free  from 
images  of  objects  not  actually  embedded  in  the  slide.  For  the  same  reason  slides 
and  covers  for  mounting  objects  to  be  photographed  must  be  cleaned  with  great 
care  and  kept  clean  until  ready  for  use.  Many  really  beautiful  sections  are  ruined 
for  photomicrographic  purposes  by  having  been  mounted  in  dirty  balsam  or  on 
dusty  slides,  or  by  being  covered  with  soiled  cover-slips.  Sections  should  be  cut  and 
mounted  in  dust-free  air,  and  the  balsam  used  in  mounting  must  be  free  from  dirt. 
Much  balsam  on  the  market  is  very  dirty  and  totally  unfit  for  mounting  sections 
designed  to  be  photographed. 

Glass  surfaces  through  which  it  is  designed  to  pass  light  should  not  be  touched 
by  the  hands,  greasy  or  otherwise.  This  applies  to  slides,  covers,  objectives,  con- 
densers, ray-filters,  photographic  lenses,  mirror  surfaces,  ground  glasses,  negatives, 
lantern-slides,  and  what  not.  The  least  touch  of  the  finger  on  a  polished  glass 
surface  generally  leaves  its  mark. 

In  using  the  common  achromatic  objectives  for  making  photomicrographs,  the 
"focus-difference"  must  be  taken  into  account.  Such  objectives,  being  corrected 
only  for  two  portions  of  the  spectrum,  require  a  different  focus  for  the  sensitive 
plate  than  for  the  human  eye.  In  other  words,  an  image  which  is  perfectly  sharp 
to  the  eye  is  not  sharp  for  the  sensitive  plate  and  will  yield  a  negative  which  is 
out  of  focus.  By  turning  the  fine  adjustment  a  measured  distance  the  image 
becomes  hazy  on  the  ground  glass,  but  will  then  yield  a  sharp  negative.  A  few 
exposures  will  determine  just  how  much  and  in  which  direction  the  eye  focus  must 
be  thrown  out  to  give  the  sharpest  result.  The  focus-difference  may  also  be  disposed 
of  by  using  monochromatic  light.  The  writer  uses  such  light  almost  altogether, 
even  with  the  best  objectives.  Another  defect  of  achromatic  objectives,  and  to  some 
extent  of  all  objectives,  is  an  arching  field,  the  center  being  out  when  the  edges 
are  in  sharp  focus.  For  this  reason  it  is  customary  to  select  a  small  portion  in  the 
center  of  the  field,  make  this  as  sharp  as  possible,  and  neglect  the  margins,  which 
may  be  trimmed  off  on  the  print.  The  Spencer  1 6-millimeter  apochromatic  objec- 
tive has  the  flattest  field  of  any  objective  of  like  quality  known  to  the  writer.  L/ack 
of  depth  of  focus  is  a  serious  defect  in  photomicrographic  work,  and  must  be  com- 
pensated for  by  making  the  sections  uniformly  thin  and  mounting  them  perfectly 
flat.  The  student  should  read  Sternberg  in  English  and  Neuhauss  in  German 
(Bibliog.,  LV). 

For  most  stained  sections  involving  bacteria,  isochromatic  plates  are  to  be 
preferred,  and  slow  rather  than  rapid  ones.  Exposure  should  be  for  contrast,  and 
consequently  as  short  as  will  give  the  necessary  detail  in  the  heavily  stained  parts. 
Development  should  be  rather  long  and  with  an  effort  to  obtain  good  contrasts. 
The  writer  formerly  used  hydrochinon,  but  now  uses  pyro,  and  develops  until  the 
image  is  visible  on  the  back. 

For  general  photographic  work  ortol  is  an  excellent  developer,  and  its  prepara- 
tion is  extremely  simple.  If  one  uses  the  Hauff  mixtures  sold  by  Gennert,  of  New 


ORTOL    DEVELOPER. 


York,  all  that  is  necessary  is  to  dissolve  one  package  of  A  in  20  ounces  of  distilled 
water  and  one  package  of  B  in  an  equal  volume  of  water  in  another  jar.  For 
normal  exposure  on  5  x  7  plates,  add  3  ounces  of  A  to  3  ounces  of  B  and  dilute  with 
2  ounces  of  water.  The  picture  begins  to  appear  in  thirty  to  forty  seconds  and 
development  is  completed  in  three  to  four  minutes.  To  soften  the  harsh  contrasts 
of  underexposed  plates  or  plates  overexposed  in  parts,  give  a  longer  development, 
using  3  ounces  of  the  alkaline  solution  to  i  ounce  or  ^  ounce  of  ortol  and  4  or  5 
of  water.  In  the  middle  of  a  long  development  it  is  often  important  to  change  to 
a  fresh  portion  of  the  developing  solution.  For  overexposed  plates,  reverse  the 
proportions,  using  i  ounce  or  l/2  ounce  of  alkali  and  3  ounces  of  the  ortol  solution 
with  several  ounces  of  water.  The  advantages  of  this  developer  are  its  quick  action 
and  its  freedom  from  stain  and  tendency  to  fog.  The  mixed  developer  may  be  used 


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over  and  over  until  exhausted  (browned).  The  quantity  named  above  will  suffice 
for  a  dozen  5x7  plates  properly  exposed.  This  developer  may  also  be  used  with 
Velox  paper.  In  this  case  it  should  be  diluted  with  more  water,  say  i  ounce  of  the 
ortol  solution,  i  ounce  of  the  alkali,  6  ounces  of  water,  and  6  drops  of  10  per  cent 
potassium-bromide  water. 

*Fic.  128.  —  Exposure  scale  set  to  show  proper  time  for  buildings  and  average  near  views  at  10 
a.  tn.  to  2  p.  m.  in  July,  with  stop  64  (32  f  )  and  an  intense  sun.  The  various  makes  of  plates  are 
divided  into  eight  classes,  and  the  time  is  read  from  the  middle  scale  for  intense  sun  and  the  most 
rapid  plates.  Under  above  conditions  a  Seed's  27-X  plate,  or  its  equivalent  (i),  would  require 
one-sixth  second.  For  light  of  a  less  degree  of  brightness  E  is  set  on  the  proper  stop,  and  the  time 
is  read  from  the  bottom  scale.  The  latter  scale  (G)  is  also  used  for  slow  plates.  With  intense 
sun,  i.  e.,  as  set  above,  a  Cramer's  isochromatic  slow  plate,  or  its  equivalent  (7),  would  require  2 
seconds.  In  indoor  work,  scale  K  is  first  set  on  H,  according  to  the  quality  of  the  light  and  num- 
ber of  windows.  Scale  L  (kind  of  walls)  is  then  set  on  the  proper  stop,  and  the  time  is  read  from 
the  bottom  scale,  according  to  the  speed  of  the  plate  used.  In  latitudes  far  to  the  north  of  Phila- 
delphia there  must  be  considerable  increase  of  time,  and  there  must  be  a  corresponding  shortening 
of  time  in  tropical  regions  or  desert  regions.  Considerable  judgment  must  also  be  used  in  making 
indoor  exposures,  especially  toward  sunset  and  soon  after  sunrise.  Near  sunset,  exposures  have 
to  be  increased  enormously.  About  three-fourths  actual  size. 


142  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

Previous  to  development  the  exposed  plate  should  be  placed  in  the  tray,  flooded 
with  water,  and  gently  rubbed  with  the  balls  of  the  fingers,  particularly  if  the 
exposures  have  been  made  for  some  time,  or  in  dusty  weather,  or  on  plates  which 
have  been  opened  for  some  time.  Many  "pin  holes"  will  be  avoided  by  this  practice, 
and  frequently  one  will  be  astonished  at  the  amount  of  dust  which  can  be  felt  as 
the  fingers  are  passed  over  the  plate. 

Negatives  should  be  fixed  in  strong  hypo  for  ten  minutes  (a  little  longer 
exposure  will  not  harm  them),  hardened  in  alum-water  (saturated)  five  or  ten 
minutes  if  the  weather  is  hot,  and  washed  in  running  water  one  to  two  hours.  If 
these  rules  are  followed,  negatives  which  are  good  on  the  start  will  not  spoil  after- 
ward. Weak  hypo  should  not  be  used,  neither  should  the  solution  be  saturated,  but 
only  nearly  so,  i.  e.,  a  saturated  solution  diluted  with  one-sixth  water.  This  is  made 
up  in  small  quantities  in  advance.  The  saturation  is  accomplished,  not  by  throwing 
the  crystals  into  a  jar  containing  water,  but  by  putting  them  into  a  cloth-sack  which 
is  brought  into  contact  only  with  the  top  layers  of  the  water.  On  removal  from 
the  washing-box  the  back  and  face  of  the  negative  should  be  rubbed  over  carefully 
under  running  tap  water  with  a  wad  of  soft  cotton,  and  set  away  in  a  clean  place 
to  dry  after  rinsing  in  distilled  water.  If  one  is  in  a  great  hurry  to  get  a  print  from 
a  wet  negative,  it  may  be  dried  in  about  ten  minutes  by  soaking  for  eight  minutes 
in  95  per  cent  alcohol  and  then  holding  it  near  an  electric  fan. 

In  developing  in  deserts  or  in  southern  climates,  in  very  hot  weather,  all  the 
fluids  must  be  iced,  including  the  wash-water,  or  else  the  plate  must  be  hardened  in 
2  per  cent  formalin  water  for  five  minutes  before  the  development  begins.  Alum- 
water  can  not  be  used  for  this  purpose,  since  it  greatly  retards  development. 

It  often  happens,  especially  with  beginners,  that  a  good  negative  (one  rightly 
exposed)  is  spoiled  by  being  left  in  the  developing  solution  too  long  or  by  being 
taken  out  too  soon.  An  overdeveloped  negative  may  be  reduced  after  soaking  it 
in  water  (or  preferably  before  it  has  dried)  by  placing  it  for  a  few  minutes  in  a  tray 
of  clean  water,  to  which  has  been  added  a  small  quantity  of  hyposulphite  of  soda 
and  a  few  drops  of  a  10  per  cent  solution  of  red  prussiate  of  potash  (Farmer's  reduc- 
ing solution),  which,  of  course,  must  be  uniformly  distributed.  Thin  negatives,  free 
from  hypo,  may  be  intensified,  if  they  are  thin  simply  from  underdevelopment,  by 
exposure  for  from  two  to  five  minutes  (occasionally  a  little  longer)  in  a  strengthening 
solution  made  of  Agfa  intensifier  20  parts  and  water  180  parts,  or  by  soaking  them 
in  a  strong  watery  solution  of  mercuric  chloride  until  they  are  whitened  through 
uniformly  on  the  back,  and  then  blacking  them  by  soaking  in  ammonia  water 
strong  enough  to  give  off  disagreeable  fumes.  If  the  time  of  exposure  is  not  nearly 
correct,  another  negative  should  be  made.  Negatives  thin  from  overexposure  do 
not  intensify  well ;  neither  do  those  which  were  much  underexposed.  All  negatives 
should  have  the  subject,  date  of  making,  and  degree  of  magnification  written  on 
them  with  a  lead  pencil  as  soon  as  they  are  dry.  The  proper  place  for  a  record  is 
on  the  margin  of  the  negative  itself  rather  than  in  a  book  or  on  a  bag.  which  may 
become  misplaced,  although  it  is  convenient  to  have  it  also  on  the  envelope,  or 
negative-bag. 


EXPOSURE-METERS.  143 

The  correct  time  of  exposure  for  photomicrographs  varies  so  greatly  with  the  size 
of  stop,  length  of  bellows,  kind  of  slide,  number  of  objective,  quality  of  light,  rapidity 
of  plate,  etc.,  that  no  very  definite  rules  can  be  laid  down,  the  right  time  in  special 
cases  in  Washington  varying  all  the  way  from  several  minutes  to  j-i_  of  a  second. 
If  the  bellows-length  is  doubled,  of  course  the  time  of  exposure  must  be  quadrupled. 
Low  powers,  and  especially  Planars,  let  through  a  great  flood  of  light  and  require 
correspondingly  short  exposures.  With  low  powers  and  sunlight  the  student  might 
begin  on  ^  second.  With  an  oil-immersion  lens  and  bright  light  he  might  try 
^  second  or  £  second.  If  the  section  is  densely  stained,  much  allowance  must 
be  made  for  that.  It  is  well,  at  least  for  a  time,  to  keep  a  record  book  of  subjects 
and  exposures  to  refresh  one's  memory.  It  saves  the  spoiling  of  many  plates.  Such 
a  record  should  include  subject,  length  of  exposure,  stop  used,  objective  and  eye- 
piece used,  length  of  bellows,  distance  of  the  condensing  lens  from  the  Abbe  con- 
denser, time  of  day,  time  of  year,  quality  of  light,  kind  of  screen,  kind  of  stain  and 
density  of  section,  kind  of  plate,  developer  used,  time  required  for  development,  and 
quality  of  negative,  viz,  overexposed,  underexposed,  or  correctly  timed. 

For  outdoor  work,  and  also  for  natural-size  or  slightly  magnified  indoor  work, 
a  good  exposure  scale  is  sometimes  useful.  The  best  ones  known  to  the  writer  are 
the  Wynne  and  the  Wager.  Success  with  the  Wynne  depends  on  one's  judgment  as 
to  the  proper  changes  in  a  good  sensitive  paper ;  with  the  Wager  it  depends  on  one's 
judgment  as  to  the  quality  of  the  light  in  the  sky.  After  a  little  experience  very 
uniform  and  excellent  results  may  be  obtained  with  either.  Personally,  the  writer 
prefers  to  use  the  Wager  (fig.  128),  because  it  is  simpler  and  takes  less  time.  No 
scale  is  always  to  be  depended  on,  there  are  so  many  variations  in  light  and  so  many 
unprovided-for  contingencies.  Experience  is  after  all  the  best  guide,  but  until  one 
has  obtained  it,  genuine  aids  are  not  to  be  neglected.  The  beginner  should  first 
become  familiar  with  the  right  exposure  for  one  stop  and  one  kind  of  plate,  e.  g., 
stop  f.  16  and  Seed's  27,  with  a  given  bellows  length.  Having  learned  correct 
exposures  under  these  constant  conditions,  it  will  be  comparatively  easy  to  change  to 
other  makes  of  plates  and  to  other  f.  stops.  Slow  isochromatic  plates  require  10  to  12 
times  as  long  exposure  as  fast  plates.  In  the  matter  ot  stops  the  length  of  exposure 
is,  of  course,  quadrupled  every  time  the  f.  stop  number  is  doubled,  and  quartered 
every  time  it  is  halved,  e.  g.,  if  stop  16  will  give  a  perfect  negative  with  one  second 
exposure,  stop  8  will  require  one-fourth  second  and  stop  32,  four  seconds.  Under 
the  same  conditions,  stop  4  will  require  one-sixteenth  second,  and  stop  64  sixteen 
seconds,  and  so  on.  With  the  Universal  stops  (those  commonly  used  on  the  shutters 
made  in  this  country  and  England)  the  exposure  is  doubled  for  the  next  higher  stop 
and  halved  for  the  next  lower  one,  instead  of  quadrupled  or  quartered,  as  in  the  case 
of  the  f.  stops. 

For  lantern  slides  the  writer  converts  a  small  room  into  a  camera  box  (plate 
1 8).  This  room  has  a  floor  space  about  6  by  5  feet.  It  has  a  north  window 
and  a  west  window.  Each  window  is  provided  with  a  double  set  of  roller  curtains, 
the  outer  made  of  yellow  cloth,  the  inner  of  a  very  dense  black  cloth  known  in  the 
trade  as  double-faced,  opaque,  black  shade-cloth,  which  lets  scarcely  any  light  through, 


144  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

even  when  held  directly  toward  the  sun.  A  cross-bar  is  screwed  across  the  base 
of  the  uprights  of  the  window  frame,  35  inches  from  the  floor  and  a  few  inches 
above  the  window  sill.  To  this  bar  a  swing  shelf  is  hinged  and  drops  down  out  of 
the  way  when  not  in  use.  This  shelf  is  about  24  inches  wide  by  30  inches  long. 
When  in  use  it  is  supported  in  a  horizontal  position  by  a  removable  leg.  On  top 
of  this  shelf  is  placed  a  cracker-box  or  some  similar  box,  to  the  sides  of  which,  at 
the  bottom,  beveled  cleats  are  nailed,  which  slide  through  corresponding  cleats 
screwed  to  the  top  of  the  shelf.  This  enables  one  to  push  the  box  toward  the  win- 
dow or  draw  it  back  on  a  regular  track.  On  the  top  of  this  box,  at  the  back  end, 
or  farthest  point  from  the  north  window,  the  camera  is  placed  facing  this  window 
and  is  screwed  fast  to  the  top  of  the  box  the  same  as  to  a  tripod.  Sidewise  move- 
ment is  provided  by  extending  the  screw-hole  in  the  top  of  the  box  into  a  slot  6  or 
8  inches  long.  In  sliding  the  camera  sidewise  it  is  of  course  necessary  to  keep  the 
ground  glass  parallel  to  the  negative  in  the  window,  and  this  is  done  by  drawing 
parallel  lines  on  top  of  the  box  about  *&  inch  apart  and  exactly  at  right  angles  to 
the  negative-carrier.  In  moving  the  camera  sidewise  all  that  then  remains  is  to 
see  that  one  side  of  the  camera  at  front  and  back  matches  one  of  these  ruled  lines. 
This  gives  ample  sidewise  movement,  and  some  up-and-down  movement  is  usually 
provided  in  the  camera  itself.  The  rest  is  obtained  by  moving  the  negative.  The 
upper  half  of  the  north  window  is  covered  by  the  curtains.  The  lower  part  is  filled 
with  a  removable  wooden  framework,  the  negative-carrier,  so  arranged  that  the 
negative  itself  may  be  moved  up  and  down  or  sidewise,  or  twisted  around  at 
will.  The  framework  of  the  negative-carrier  is  made  of  inch  stuff.  When  in  use 
it  is  placed  upright  about  3  inches  in  front  of  the  window  pane  and  just  behind 
the  cross-bar  which  keeps  it  in  place.  In  the  middle  of  this  frame  is  a  circular 
wooden  disc  (which  must  turn  freely),  held  in  place  on  the  back  by  a  ledge  and  in 
front  by  four  buttons,  and  open  in  the  center.  The  breadth  of  this  disc  is  24  inches, 
and  it  should  be  made  of  well-seasoned  lumber  of  a  sort  not  inclined  to  warp.  On 
this  disc  at  either  side  two  broad  vertical  cleats  are  fastened.  These  are  grooved 
on  the  inner  edges  next  the  framework,  and  under  them,  close  to  the  circular  piece, 
two  wide  ^4-inch  pieces  slide  up  and  down  freely,  carrying  the  negative  between 
them.  The  latter  fits  into  marginal  grooves  and  is  held  in  place  by  buttons.  The 
marginal  grooves  extend  the  whole  length  of  the  ^-inch  pieces  and  consequently 
allow  the  negative  to  be  moved  sidewise  to  any  extent  desired,  while  the  up-and- 
down  movement  is  obtained  by  sliding  the  two  ^-inch  pieces  and  the  negative 
between  them  as  a  unit.  Over  the  first  grooved  cleats,  at  right  angles,  i.  e.,  horizon- 
tally, two  similar  cleats  are  screwed.  These  also  have  wide  s^-inch  pieces  moving 
under  their  grooved  edges.  These  sliding  pieces  cover  the  sides  of  the  negative 
and  shut  out  the  side-light  in  whatever  position  the  negative  may  be  placed.  Behind 
the  negative  against  the  window  is  pasted  (by  its  corners)  a  good-sized  piece  of 
white  tissue-paper,  which  serves  to  distribute  the  light  evenly  and  to  cut  out  images 
of  trees,  buildings,  etc.  When  in  use  the  double  curtains  of  the  west  window  are 
drawn  down  and  the  door  is  shut.  The  north  light  which  enters  the  room  then 
comes  through  the  negative  placed  in  front  of  the  camera.  The  focus  is  obtained 


PLATE  18. 


Small  room  arranged  for  making  lantern  slides  and  enlargements  on  bromide  paper. 

The  parts  of  the  window  shutter  are  as  follows :  ( I )  frame  work,  (2)  circular  piece  giving  rotary  motion,  (3)  one  ol  Iwo  stationary  piece» 
under  which  No.  4  slides,  (4)  negative  carrier,  ( 5)  one  of  two  stationary  pieces  under  which  No.  6  slides,  (6)  side  pieces  designed  to  cut 
out  all  the  side  light  except  that  which  comes  through  the  negative. 


PHOTOGRAPHY   AND    PHOTOMICROGRAPHY. 


145 


on  the  ground  glass  as  for  any  picture,  remembering  that  a  wide  margin  (^  inch  or 
more)  must  be  left  for  binding  strips,  and  that  if  the  negative  has  any  up  and  down 
its  image  must  be  placed  crosswise  on  the  lantern-slide.  The  writer  focuses  as 


Fig.  129.* 


*Fic.  129.— The  modified  Collins-Brown  camera  used  with  tripod  for  natural-size  pictures. 
Heavy  shadows  are  dissipated  by  using  the  glass  plate.  The  size  of  the  camera  box  is  IO}4  by  I2J4 
by  SJ4  inches,  and  its  weight,  including  lens  and  shutter,  is  about  15  pounds,  or  with  tripod  19 
pounds.  The  camera  takes  a  6j4  by  8J4  plate.  It  is  solidly  constructed,  of  the  very  best  workman- 
ship, and  the  only  objection  is  its  weight,  which  is  no  disadvantage  in  laboratory  use.  It  is  not 
recommended  for  field  use. 


146 


BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 


Fig.  130* 


sharp  as  possible  with  stop  wide  open  and  then  stops  down  to  16  u.  s.  before  making 
the  picture. 

There  are  two  other  ways  of  making  lantern-slides,  i.  <?.,  by  contact  exposure, 
the  gelatin  films  face  to  face,  and  by  means  of  a  long  box-camera  with  the  negative 
in  one  end,  the  lantern-slide  carrier  in  the  other  end,  and  the  lens  between  the  two,  i.  e., 
inside  the  camera-box,  held  in  a  framework  sliding  between  the  two  ends  and  having 

front  and  rear  bellows  attached  to  its  outside  parts. 
The  method  by  contact  exposure  is  not  very  satis- 
factory unless  the  negative  and  the  lantern-slide  are 
of  the  same  size.  The  box-method  is  a  very  good 
one.  A  box  of  this  kind  is  very  convenient,  and 
may  also  be  used  for  enlargements  up  to  5  or  6 
times.  The  bellows-extension  should  be  ample,  so 
I  that  various  lenses  may  be  accommodated  and  so 
I  that  lantern-slides  maybe  made  from  large  negatives 
if  desired,  i.  <?.,  the  solid  framework  or  track  on 
which  the  parts  slide  should  be  about  6  feet  long, 
and  the  bellows  -extension  to  either  side  of  the 
middle  piece  should  be  not  less  than  3  feet,  exclusive  of  the  woodwork  at  each  end 
and  in  the  middle.  A  very  good  apparatus  of  this  sort  is  shown  in  plate  17.  It  is 
the  Folmer  &  Schwing  enlarging,  reducing,  and  copying  camera, 
mounted  on  a  plain  wooden  table  of  home  construction,  and  the 
only  defect  I  have  discovered  in  it  is  that  it  has  too  short  a  bellows 
for  use  with  lenses  having  a  1  2-inch  focus.  It  has  a  very  neat  device 
for  obtaining  a  sharp  focus  and  many  other  conveniences,  and  might 
just  as  well  be  made  with  a  longer  bellows.  It  is  convenient  to 
have  a  box  which  will  take  1  1  by  14  plates.  When  making  lantern- 
slides  the  end  of  the  box  carrying  the  negative  is  pointed  toward 
the  window  and  is  elevated  a  foot  or  more  to  secure  uniform  light- 
ing. The  writer  has  found  the  Voigtlaender  collinear  lens,  series  III, 
No.  6,  very  satisfactory  for  making  lantern-slides  and  enlargements. 
In  plate  17  the  bellows-extension  used  when  making  lantern-slides 
from  large  negatives  lies  on  the  floor. 

The  time  of  exposure  for  lantern-slides  varies  greatly  with  quality 
of  light  and  density  of  negatives,  e.  g.,  with  stop  64  u.  s.  from 
YZ  second  or  less  in  bright  light  to  fifteen  minutes  or  more  in  very 
dull  light  with  dense  negatives.  L,antern-slides  should  not  be  developed  with  pyro 
because  it  stains,  and  should  not  be  developed  with  metol-hydro  because  it  often 
gives  a  foggy  appearance  if  the  contrasts  in  the  negative  are  great  or  the  exposure 
is  a  little  too  long.  Hydrochinon  gives  very  satisfactory  slides. 


*'       * 


*FiG.   130. — Cross-level,  made  by  The  L.  S.  Starrett    Company,   Athol,    Mass.     Nearly   actual 
size.    This  is  very  convenient  for  use  with  cameras. 

fFio.  131. — Device  for  cutting  out  light  in  air-shaft  of  dark-room.    Diameter,  12  inches. 


LANTERN    SLIDES,    ETC. 


147 


In  lantern-slides  one  desires  much  detail  and  little  density ;  it  is  customary, 
therefore,  to  develop  only  until  there  is  a  good  surface  image.  On  no  account  must 
the  development  be  pushed  until  the  image  shows  through  on  the  back.  Even  the 
densest  portions  must  be  translucent.  Slides  suitable  for  projection  with  very  bright 
lights  may  prove  too  dense  for  dull  ones.  In  making  lantern-slides  one  must  keep 
in  mind  the  kind  of  light  to  be  used  in  projection. 

In  making  enlargements,  the  camera  is  lowered  to  the  horizontal  and  pointed 
away  from  the  window,  and  the  object  is  lighted  in  part  from  above  (skylight).  To 


Fig.  132* 

facilitate  shifts  it  is  convenient  to  have  the  table-top  (to  which  the  camera-bed  is 
screwed)  turn  freely  on  a  central  pivot,  and  the  legs  of  the  table  should  be  mounted 
on  casters  so  that  it  may  be  moved  about  easily.  The  table  devised  by  the  writer  is 
shown  in  plate  17.  This  was  built  by  a  carpenter  and  does  well  enough.  In 
making  enlargements  the  lens-board  is  removed  from  the  interior  and  substituted 
for  the  kits  in  the  front  end  of  the  camera  ;  the  ends  of  the  wooden  carrier  (shown 
on  first  shelf  of  the  camera-table)  are  slid  under  the  beveled  cleats  at  the  front  end 
*Fic.  132.— Side  view  of  a  convenient  small  dark-room,  devised  by  Mr.  Hubbard. 


148 


BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 


of  the  table,  and  the  map  or  other  object  to  be  enlarged  is  then  pinned  on  a  flat 
board  in  the  right  position  in  front  of  the  lens,  the  board  being  held  in  place  by  the 


SHELVES  FOR 
STOCK  DCVCLOPCR 

WASHING 

CUPBOARD 

I 

DE.VEL         FIXIN 

3          ALUM 

E 

ll         : 

Q 
C 

SHELVES 

" 

I 

Q 


STOCK  BOTTLES 
OF  DEVELOP/KG  SOLUTIONS 


STOCK  BOTTLES 
OF  DEVELOPING  SOLUTIONS 


"'IT0 

WATLR 


fXTRA   TRAYS 


BROMIDE  SOL 


k /oft.      

Fig.  133* 

carrier.     The  desired  magnification  is  obtained  by  sliding  the  carrier  in  or  out  to  a 
marked  place  previously  determined. 

When  not  made  directly  from  the  micro- 
scope, the  histological  drawings  in  this  book 
have  been  made  from  photographic  enlarge- 
ments.    For  example,  in  fig.  72  asolio  print 
or  bromide  print  was  made  from  the  photo- 
micrograph.    This  was  then  enlarged  three 
or  four  times  and  from  the  resulting  nega- 
tive a  salted-paper  silver  print  or  a  blue  print 
was  made.     A  drawing  was  then  made  on 
this  print  with  a  fine-pointed  pen  and  water- 
proof India  ink.    After  careful  inspection  by 
the   writer   and  such  changes  as   were   re- 
quired to  make  the  drawing  correspond  more 
nearly  in  all  its  details  to  the  main  lines  of 
the  photograph,  the  brown  of  the  silver,  or 
the  blue  of  the  iron  salt,  was  removed  by  a 
bath  in  water  containing  cyanide  of  potash. 
On  reduction  by  the  photoengraver  many  of 
the  inequalities  in  the   pen-work   of  such 
drawings  disappear  and  the  pictures  closely 
resemble  the  originals,  whereas  if  they  are 
drawn  without  enlargement,  and  engraved 
as  drawn,  the  pen  lines  will  in  many  cases 
have  a  more  or  less  ragged  appearance. 

*Fic.  133. — Top  view  of  the  room  shown  in  fig.  132.    All  of  the  trays  rest  on  triangular  slats 
covering  a  deep  sink.    The  screens  are  raised  and  lowered  very  easily  by  balanced  weights. 
j-Fii.  134. — Side  view  of  a  small  photographic  dark-room  in  Laboratory  of  Plant  Pathology. 


MIXED 
DCfCLOPCK 


HYPO    SHELF 


STOCK  BOTTLES 
OFHYPO  AND  ALUM 


Fig.  I34.f 


PLATE  19. 


Bacterial  black  spot  of  the  plum. 

Spots  about  six  weeks  old,  except  •  very  few  on  the  two  right-hand  plums  of  the  upper  row.  Fruits  about  one-halt  grown  ;  collected 
July  24,  1902,  from  an  orchard  of  Japaneie  plunu  in  central  Michigan.  In  this  stage  the  plums  (var.  Hale)  begin  to  crack  open 
and  are  attacked  occasionally  by  fungi  (MoniHa,  etc.).  Some  tufts  of  Monilia  may  be  seen  on  the  outer,  left-hand  fruit,  second 
row  from  the  bottom.  Early  stages  of  this  disease  are  shown  in  figs.  70,  7 1 ,  and  72. 


VENTILATION    OF   DARK-ROOM. 


149 


LOADING     SHELF 


f  ft.  9  in. 


DOOR  \ 

Fig.  135.* 

in  use  much  of  the  time,  some 
The  writer  accomplishes  this 
by  an  electric  fan  placed  in  the 
mouth  of  an  air-shaft  which 
extends  from  the  ceiling  to  6  or 
8  feet  above  the  roof.  These 
shafts  are  cylindrical,  i  foot  in 
diameter,  made  of  heavy  sheet- 
iron  and  surmounted  by  a  broad, 
mushroom  -  shaped  cap.  The 
interior  is  painted  a  dead  black, 
and  as  an  additional  precaution 
against  the  entrance  of  light  it 
carries  a  sleeve  of  the  fonn 
shown  in  fig.  131.  This  effectu- 
ally cuts  out  light.  The  air  is 
pumped  out  so  rapidly  by  a  de- 
vice of  this  sort  that  not  the 
least  inconvenience  is  experi- 
enced in  working  all  day  in  a 
very  small  room. 

If  only  one  or  two  persons 


The  very  convenient  heavy  camera 
shown  in  fig.  124  may  also  be  used 
for  natural-size  work,  arranged  as 
shown  in  fig.  129.  In  this  connection 
the  Starrett  cross-level  shown  in  fig. 
130  will  be  found  very  convenient 
for  leveling  the  back  of  the  camera. 

Very  excellent  cameras  are  made 
by  the  Century  Camera  Company. 
Their  Long-focus  Century  Grand 
leaves  little  to  be  desired  in  the  way 
of  a  convenient,  perfect-working  in- 
strument. 

The  dark-room  for  development  is 
an  important  subject.  The  chamber 
must  be  light-tight.  At  the  same 
time  it  ought  to  be  roomy  and  well 
ventilated.  If  the  room  is  small  and 
means  of  removing  the  foul  air  becomes  imperative. 


5; 
o 


I 


Fig.  I36.t 


*Fic.  135. — Diagram  of  small  dark-room  shown  in  fig.  134.  Standing  in  the  middle,  a  man  can 
touch  the  walls  in  either  direction.  In  the  ceiling  is  a  foot-wide  pipe  extending  6  feet  beyond  the 
roof  and  capped  with  a  broad  mushroom  top.  In  the  lower  end  of  this  pipe  is  an  electric  fan, 
which  pumps  foul  air  out  of  the  room.  Light  is  prevented  from  entering  by  partial  cross-septa 
projecting  from  opposite  sides  of  the  air-shaft,  and  also  by  blacking  its  inner  surface. 

fFic.  136. — Wall  case  for  preserving  from  dust  and  scratches  the  enameled  iron  plates  used  for 
squeegeeing  silver  prints. 


150  BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 

of  good  habits  are  to  use  the  dark-room,  it  is  very  convenient  to  have  developing 
dishes  and  fixing  trays  on  the  same  shelf,  which  may  be  of  slats  over  a  deep  sink, 
as  in  figs.  132  and  133.  With  some  shelves  over  this  sink  and  a  water-tap  above  it, 
everything  is  in  reach  without  moving  about.  If,  on  the  contrary,  various  persons 
are  to  work  in  the  dark-room,  some  of  them  students  with  unformed  habits,  some 
of  them  older  workers  with  incorrigibly  slovenly  habits,  including  a  disposition  to 
spill  hypo  over  everything,  then  some  different  arrangement  must  be  made,  the 
sodium  hyposulphite  trays  especially  being  kept  on  a  separate  shelf  at  a  good  distance 
from  the  developing  shelf.  Figs.  134  and  135  show  the  arrangement  of  a  small  dark- 
room devised  by  the  writer  for  photographic  work,  the  space  at  his  disposal  being 
very  limited.  The  air-shaft  is  in  the  ceiling  over  the  loading  shelf.  Artificial  light 
is  furnished  by  two  i6-candle  power  Edison  electric-light  bulbs,  one  hanging  near 
the  wall  above  the  sink,  the  other  inclosed  in  the  red-light  box.  Over  the  ruby 
glass  there  is  placed  a  sheet  of  orange-buff  paper,  commonly  called  post-office  paper. 
The  hypo  trays  are  under  the  sink.  The  zinc  box  for  washing  negatives  stands  in 
the  sink.  Developers  are  mixed  on  the  drop-shelf,  and  are  kept  on  the  shelves 
above  it  The  bromide  bottle,  graduates,  and  beakers  are  kept  on  the  small  shelves 
above  the  developing  shelf.  Large  bottles  of  alum  solution,  hypo  solution,  etc.,  are 
stored  under  the  hypo  shelf. 

Enameled  plates  for  squeegeeing  silver  prints  may  be  stored  when  not  in  use  as 
shown  in  fig.  136.  In  this  way  they  are  protected  from  dust  and  scratches.  To 
prevent  prints  from  sticking  the  surface  of  the  plates  is  occasionally  nibbed  with 
paraffin  dissolved  in  xylol,  and  is  then  polished  with  a  soft  clean  cloth. 

Some  memoranda  on  photographic  developers  will  be  found  under  Formula. 
In  addition  to  what  has  been  said  there  a  few  hints  on  salted  silver-paper,  blue- 
print paper,  and  bromide-papers  may  be  of  service  to  those  who  wish  to  use  these 
methods  as  preliminary  to  pen-and-ink  drawing. 

All  papers  designed  for  this  use  should  possess  a  smooth  surface  suitable  for 
pen-and-ink  work,  arid  sensitive  papers  of  this  quality  may  be  had  of  various  dealers 
by  specifying  just  what  is  desired.  Blue-print  paper  and  salted  silver-paper  may  be 
made  for  one's  self.  It  is  preferable,  however,  to  purchase  the  former,  and  to  make 
the  latter  or  to  buy  it  fresh,  as  it  does  not  keep  well.  Directions  for  making  the 
plain  silver-paper  may  be  found  in  "  The  Figures,  Facts,  and  Formulae  of  Photog- 
raphy and  Guide  to  their  Practical  Use,"  by  H.  S.  Ward,  N.  Y.,  Tennant  &  Ward, 
1903;  and  in  "The  Photo  Miniature,"  Vol.  II,  No.  22,  "Albumen  and  Plain  Paper 
Printing." 

All  sensitive  papers  placed  in  the  printing  frame  should  have  the  coated  side 
next  to  the  film-side  of  the  negative. 

Blue-print  paper  is  much  less  sensitive  to  light  than  solio  paper,  and  long  solar 
printings  are  required.  When  the  paper  has  assumed  a  deep-bronzy  appearance  it 
may  be  assumed  to  be  sufficiently  printed.  A  few  trials  will  give  the  necessary 
experience.  Blue-prints  are  developed  by  simply  washing  them  in  several  changes 
of  pure  water,  taking  care  that  the  coated  surface  is  wetted  thoroughly  from  the 
start,  i.  e.,  freed  from  air-bubbles.  No  fixing  is  required. 


PLATE  20. 


Bacterial  disease  of  broomcorn.    (Hothouse,  U.  S.  Department  of  Agriculture,  spring  of  1905.) 

Disease  not  complicated  by  aphides.  The  signs  consist  of  gradually  elongating  red  or  red-brown  stripes  on  a  green  background.  In  later  stages  the  stripes  coalesce 
and  the  leaves  shrivel.  Under  surface  of  diseased  areas  often  covered  by  red  crusts — the  dried-down,  bacterial  ooze  from  interior  of  leaves.  The  infections  were 
by  way  of  the  stomaU  and  these  spots  are  about  six  weeks  old. 


DRAWINGS   ON    PHOTOGRAPHIC    PRINTS. 

Salted  silver-paper  also  requires  a  rather  long  printing  with  sunlight.  When 
properly  printed  the  paper  is  washed  in  a  bath  of  salt-water,  rinsed  in  several 
changes  of  pure  water,  and  fixed  in  a  weak  solution  of  hypo  (i :  15). 

Bromide-prints  under  the  negative  are  usually  made  by  exposing  the  paper 
to  artificial  light  at  a  standard  distance,  say  9  inches  or  12  inches.  By  always 
exposing  at  a  given  distance  and  to  a  light  of  uniform  intensity  two  variable  factors 
are  excluded,  and  one  then  has  to  take  into  account  only  the  quality  of  the  paper 
and  the  density  of  the  negative.  It  is  usually  economical,  especially  for  beginners, 
to  test  the  density  of  the  negative  in  advance  by  exposing,  for  various  periods, 
narrow  strips  of  the  sensitive  paper  laid  across  the  negative  so  as  to  include  dense 
and  thin  portions.  These  strips  are  then  developed,  and  if  none  of  them  have  been 
properly  exposed,  a  second  trial  is  made.  When  the  right  exposure  has  been 
learned,  the  print  is  made.  A  little  experience  enables  one  to  judge  quite  correctly 
as  to  the  proper  exposure  by  simply  looking  through  a  negative.  No  very  definite 
rules  can  be  given  for  length  of  exposure ;  this  depends  so  much  on  distance  from 
the  light  and  brightness  of  the  flame.  With  velox  paper  and  an  ordinary  flat  gas- 
flame  at  a  distance  of  9  inches  the  writer's  negatives  usually  require  from  15  seconds 
to  2  minutes.  With  a  Welsbach  light  or  with  thin  negatives  the  time  would  be 
shorter.  At  a  distance  of  18  inches  from  the  light  the  time  would,  of  course,  be 
quadrupled.  Directions  for  the  employment  of  special  developers  usually  accom- 
pany each  maker's  paper.  The  writer  has  found  ortol  (p.  141)  a  very  good  developer 
for  velox  paper,  and  prefers  it  to  metol-hydro.  Velox-prints  are  developed  in  weak 
artificial  light  (gas  turned  low) ;  they  are  rinsed  from  the  developing  solution  by 
passing  quickly  through  a  bath  of  pure  water ;  they  are  then  fixed  in  hypo,  and 
washed  for  at  least  one-half  hour  in  running  water.  The  writer  pins  the  prints  to 
a  smooth  board  and  floats  these  in  a  bath-tub  or  clean  sink,  film-side  down.  Most 
of  the  curling  may  be  avoided  by  drying  the  prints,  film-side  down,  on  mosquito- 
netting  stretched  on  a  wooden-frame. 

The  yellowing  of  prints  is  often  due  to  the  fact  that  they  were  not  properly 
fixed;  the  hypo  solution  was  weak,  or  the  time  of  exposure  to  it  was  not  sufficient. 

All  pen-and-ink  drawings  on  such  photographic  prints  must  be  made  with 
waterproof  India  ink,  after  which  the  photographic  part  is  bleached  out  by  exposure 
for  a  few  minutes  in  water  containing  cyanide  of  potash  (i  1500,  more  or  less).  The 
drawings  should  be  exposed  in  this  bath  only  as  long  as  necessary.  If  any  part  of 
the  print  refuses  to  bleach,  it  should  be  moistened  with  iodine  potassium  iodide  and 
returned  to  the  cyanide  bath.  It  is  then  passed  through  pure  water  and  dried  face 
up  on  blotting  paper  in  a  place  free  from  dust. 

SOME   MILESTONES   IN   THE   PROGRESS   OF   BACTERIOLOGY. 

The  development  of  bacteriology  can  not  be  separated  from  advances  in  human 
and  animal  pathology.  Physicians  and  surgeons  have  made  most  of  the  brilliant 
discoveries  or  have  led  the  way  to  them.  Chemists  and  physicists  have  assisted. 
With  a  few  shining  exceptions,  botanists  have  had  comparatively  little  to  do  with 
the  advancement  of  this  science.  Bacteriological  inquiry  has  been  an  incentive  to 


152  BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 

the  improvement  of  various  kinds  of  apparatus,  notably  the  microscope,  and  it  has 
derived  corresponding  advantages  from  the  use  of  these  improved  instruments  of 
research.  We  owe,  in  particular,  a  great  debt  to  the  German  physicist  Abbe,  whose 
discovery  of  the  Jena  glass  made  possible  the  superb  modern  apochromatic  objective. 
Among  the  multitude  of  workers  in  animal  pathology  and  bacteriology  during 
the  last  thirty-five  years  certain  men  tower  far  above  the  rest,  their  contributions  to 
science  having  been  more  conspicuous  and  their  imprint  on  their  generation  more 
lasting.  If  France  is  mentioned,  we  think  at  once  of  Pasteur,  Davaine,  Duclaux, 
Metchnikoff,  Chamberland,  Roux,  Nocard,  and  Chauveau.  In  Germany  we  think 
of  Virchow,  Cohn,  Cohnheim,  Koch,  Weigert,  Nicolaier,  Eberth,  Gaffky,  Hueppe, 
Fliigge,  Fraenkel,  Pfeiffer,  Behring,  Ehrlich,  and  many  others ;  in  Japan,  of  Kitasato 
and  Shiga;  in  the  United  States,  of  Welch,  Sternberg,  Theobald  Smith,  Nuttall, 
Councilman,  and  a  host  of  brilliant  younger  men,  many  of  whom  received  their 
training  under  Welch  in  the  Johns  Hopkins  Pathological  Laboratory.  England, 
from  which  one  might  have  expected  so  much,  has  contributed  comparatively  little, 
owing  probably  to  the  laws  in  force  in  that  country  respecting  animal  experimenta- 
tion, laws  framed  with  the  intention  of  doing  a  kindness  to  the  lower  animals,  but 
working,  on  account  of  their  interference  with  the  pathologist,  a  distinct  detriment 
both  to  men  and  animals,  the  aim  of  all  animal  pathological  inquiry  being  the  alle- 
viation of  human  and  animal  suffering.  In  passing  we  should  not  forget,  however, 
the  contributions  of  Tyndall  and  Lister,  the  one  a  physicist,  the  other  a  surgeon. 

.  Undoubtedly  bacteriology  owes  very  much  to  Louis  Pasteur.  France  has  had 
many  great  sons,  none  greater  than  he.  His  refutation  of  the  doctrine  of  spontane- 
ous generation  cleared  the  air  of  many  misconceptions  and  laid  the  foundations  for 
exact  experimentation.  His  demonstration  of  the  nature  of  pebrine  and  flacherie, 
two  destructive  diseases  of  silk  worms,  brought  again  into  vivid  light  the  assump- 
tion that  the  origin  of  a  great  variety  of  human  and  animal  diseases  should  be 
sought  in  the  activities  of  microscopic  organisms.  His  studies  of  anthrax  and 
other  diseases  of  warm-blooded  animals  confirmed  this  suspicion  and  set  a  great 
many  persons  thinking  and  working.  His  investigations  of  the  problems  connected 
with  fermentation  were  similarly  fertile  in  discovery  and  in  suggestion. 

The  publication  of  Robert  Koch's  great  paper  on  tuberculosis  in  1884  marked 
another  distinct  advance.  The  same  memorable  year  Koch  published  in  full  his 
discovery  of  the  cause  of  Asiatic  cholera,  only  a  brief  announcement  of  it  having 
been  made  in  1883.  The  whole  world  was  interested,  and  from  this  time  on  experi- 
menters began  to  multiply  in  every  civilized  land,  boards  of  health,  universities,  and 
private  citizens  vying  with  each  other  in  the  establishment  of  laboratories  for  the 
study  of  these  minute  organisms  endowed  with  so  much  power  for  good  or  evil. 
Koch's  investigations  in  South  Africa  bring  us  down  to  recent  times,  where  the  per- 
spective is  not  so  good.  To  sum  up  very  briefly,  omitting  many  things,  the  following 
are  some  of  the  milestones : 

i.  Overthrow  of  the  doctrine  of  spontaneous  generation. 

,;  ,  ,  2.  Discovery  that  putrescible  fluids  (exclusive  of  milk)  will  not  decay  after 
boiling,  if  protected  from  the  bacteria  of  the  air  by  means  of  cotton-plugs. 


MILESTONES.  153 

3.  Pasteur's  studies  of  fermentations ;  discovery  of  anaerobic  organisms. 

4.  Pasteur's  studies  of  p£brine,  flacherie,  anthrax,  chicken-cholera,  and  rabies. 

5.  Cohn's  system  of  classification. 

6.  Cohn's  discovery  of  endospore-bearing  organisms  resistant  to  heat. 

7.  Introduction  of  anilin  stains  and  photomicrography. 

8.  Tyndall's  discontinuous  moist  sterilization. 

9.  Lister's  antiseptic  surgery. 

10.  Lister's  dilution  method  for  obtaining  pure  cultures.     Ascribed  also  to 
Naegeli. 

n.  Miquel's  discovery  of  thermophilic  bacteria. 

12.  Discovery  of  root-tubercles  of  Leguminosse  by  Woronin,  and  subsequent 
papers  by  Beyerinck,  Hellriegel  &  Wilfarth,  et  al. 

13.  Discovery  of  bacterial  diseases  in  plants  by  Burrill,  Prillieux,  and  Wakker. 

14.  General  introduction  of  Koch's  poured-plate    method  for  obtaining  pure 
cultures. 

15.  Koch's  discovery  of  the  "comma  bacillus,"  the  cause  of  Asiatic  cholera. 

1 6.  Paper  on  tuberculosis  by  Koch  in  Mitth.  a.  d.  Kaiserlichen  Gesundheitsamt, 
Bd.  II. 

17.  Use  of  synthetic  media,  of  pressure  filters,  of  fermentation  tubes,  and  of 
other  anaerobic  apparatus. 

1 8.  Introduction  of  apochromatic  objectives. 

19.  Eberth  and  Gaffky's  discovery  of  the  bacillus  of  typhoid  fever  ;  Nicolaier's 
discovery  of  the  tetanus  bacillus ;  LoefHer  &  Schutz's  discovery  of  the  bacillus  of 
glanders ;  Salmon  &  Smith's  discovery  of  the  hog-cholera  bacillus ;  Yersin's  and 
Kitasato's  independent  discovery  of  the  bacillus  of  plague ;   Pfeiffer's  discovery  of 
the  organism  causing  influenza ;  Shiga's  discovery  of  the  cause  of  tropical  dysentery. 

20.  Winogradsky's  studies  of  nitrifying  organisms. 

21.  Hansen's  studies  of  fermentation,  more  especially  yeasts. 

22.  Duclaux's,  Greene's,  and  Brown  &  Morris's  study  of  enzymic  actions. 

23.  Study  of  toxines  and  anti-toxines ;  general  use  of  anti-diphtheritic  serum. 

24.  Migula's  attempt  to  determine  the  exact  morphology  of  all  known  species. 

25.  Discovery  of  the  cause  of  peripneumonia  in  cattle  by  Nocard  &  Roux; 
organism  so  minute  as  to  be  at  the  extreme  limit  of  microscopic  definition. 

26.  Profound  specialization,  resulting  in  distinct  classes  of  bacteriologists,  e.  g., 
animal  and  plant  bacteriologists,   milk  bacteriologists,  water  bacteriologists,  soil 
bacteriologists;  and  in  special  societies  and  journals,  e.g.,  those  devoted  exclusively 
to  the  study  of  tuberculosis. 

Beyond  this  field,  but  of  extreme  pathological  interest,  and  worked  out  by  the 
exact  methods  of  the  bacteriologist,  are  Laveran's  discovery  of  the  protozoan  causing 
malarial  fevers  and  Theobald  Smith's  discovery  of  the  protozoan  causing  the  bovine 
disease  known  as  Texas  fever,  both  parasites  of  the  red  blood-corpuscles.  More 
recently  it  has  developed  that  these  are  not  rare  types  of  disease.  On  the  contrary, 
many  virulent  diseases  of  man  and  the  lower  animals  are  now  known  or  believed 
to  be  due  to  Tripanosomes  or  other  Protozoans,  and  the  literature  on  the  subject 
is  becoming  voluminous. 


154  BACTERIA  IN  RELATION  TO  PLANT  DISEASES. 

NOMENCLATURE  AND  CLASSIFICATIONS. 

The  nomenclature  of  the  bacteria  is  in  a  somewhat  chaotic  state,  as  might  be 
expected  of  a  science  which  has  been  cultivated  so  largely  by  medical  men  and  so 
comparatively  little  by  systematic  botanists  and  zoologists.  The  writer  therefore 
will  venture  a  few  remarks  on  this  subject. 

If  an  organism  is  distinct  from  any  which  has  been  described,  so  as  to  be 
regarded  as  a  new  species  or  spoken  about  as  a  distinct  thing,  then  it  should  be  given 
a  specific  L/atin  name  and  not  designated  by  a  figure  or  a  letter  of  the  alphabet. 
Bacillus  No.  i,  2,  or  3,  or  A,  B,  C,  is  proper  enough  for  private  memoranda  while 
an  investigation  is  incomplete,  but  when  it  is  finished  and  ready  for  publication 
these  designations  should  give  place  to  scientific  names. 

Naturalists  everywhere  are  in  agreement  that  the  scientific  name  of  a  living 
thing  should  consist  of  two  words  only — the  name  of  the  genus,  followed  by  the 
name  of  the  species,  after  which  is  usually  added  the  name  of  the  author,  or,  if  a  trans- 
position has  been  made  from  one  genus  to  another,  the  name  of  the  original  describer 
is  put  into  a  parenthesis,  followed  by  that  of  the  transferrer  outside  of  the  parenthesis. 
All  polynomials,  of  which  there  are  now  many,  are  to  be  regarded  as  nomina  exclu- 
denda.  For  example,  Bacillus  coli  communis  should  give  place  to  B.  eoh\  and  such 
names  as  Bacillus  membranaceus  amethystinus  mobilis,  Bacillus  argenieo  phosphor- 
escens  liquefaciens,  Bacillus  pyogenes  foetidus  liquefaciens,  should  yield  to  something 
shorter  and  more  in  conformity  with  modern  views  of  nomenclature.  More  than 
170  trinomial  names  are  to  be  found  in  the  last  edition  of  Fliigge's  Mikroorganis- 
men,  and  very  few,  if  any,  of  them  were  given  with  the  distinct  idea  that  they  repre- 
sent varieties  of  other  organisms.  The  habit  of  giving  trinomial  or  quadrinomial 
names  should  be  abandoned,  and  as  far  as  possible  binomial  names  should  be  sub- 
stituted for  those  already  in  literature. 

In  the  period  antedating  Koch's  discovery  of  the  poured-plate  method,  when 
there  was  no  very  satisfactory  way  of  separating  one  organism  from  another  so  as 
to  have  pure  cultures,  the  descriptions  were  necessarily  vague.  They  were  usually 
drawn  from  mixed  cultures,  and  very  brief  descriptions  were  supposed  to  be  ample. 
The  result  is  that  many  of  the  names  which  have  come  down  from  this  period  are 
nomina  nuda,  or  semi-nuda,  i.  e.,  it  is  impossible  to  associate  them  with  any  known 
organism  for  the  very  good  reason  that  they  were  not  founded  on  any  one  organism, 
but  on  mixtures  now  indeterminable,  or  were  too  imperfectly  characterized.  Gen- 
erally speaking,  such  names  should  be  abandoned.  The  only  safe  rule  and  the  only 
just  one  is  to  discard  all  specific  names  which  do  not  carry  with  them  an  exact 
statement  or  description,  sufficient  to  associate  the  name  beyond  doubt  with  a  par- 
ticular organism.  It  is  not  sufficient  description  of  an  organism  to  say  that  it  is 
the  cause  of  a  disease,  unless  the  author  has  proved  it  to  be  such  according  to  the 
well-recognized  rules  of  pathology.  In  order  that  his  name  shall  hold,  an  author 
must  have  carefully  described  the  disease  and  must  have  proved  in  some  way  the 
pathogenic  nature  of  his  organism,  or  else  he  must  have  given  a  fairly  correct  descrip- 
tion of  the  morphology  and  physiology  (cultural  characters)  of  the  organism,  so  that 
it  can  be  detected  anywhere.  Just  how  much  shall  constitute  a  sufficient  description 
must  depend  on  circumstances.  A  few  lines  might  be  sufficient  if  the  description 


NOMENCLATURE   AND    CLASSIFICATIONS.  155 

were  exact  and  of  such  a  character  as  to  definitely  indicate  a  particular  organism 
as  the  one  intended.  Many  pages  would  be  insufficient  if  the  description  is  vague 
and  contradictory  and  does  not  enable  the  scientific  public  to  fix  upon  a  particular 
organism  as  the  one  intended.  The  careful  work  of  subsequent  investigators  may 
sometimes  lead  an  author  to  say  that  he  meant  to  designate  such  or  such  organisms 
by  his  names,  but  if  he  really  described  something  different  or  made  no  intelligible 
descriptions,  then  his  names  can  only  be  regarded  as  equivalent  to  nomina  nuda  and 
should  never  be  substituted  for  later  ones  given  after  careful  study  and  description 
of  the  organism.  Any  other  course  puts  a  premium  on  bad  work.  In  case  of  the 
higher  plants  and  animals,  preserved  specimens  will  often  serve  to  correct  a  faulty 
description  and  to  indicate  clearly  the  object  to  which  the  name  was  applied.  Cultures 
of  particular  bacteria  kept  alive  by  means  of  frequent  transfers  to  fresh  culture-media 
will  also  serve  the  same  purpose  when  they  are  able  to  run  the  gauntlet  of  extermina- 
tion by  other  organisms  accidentally  introduced  during  some  one  of  the  many  trans- 
fers, and  when  they  have  not  varied  too  greatly  from  the  original  type  as  a  result  of 
changed  environment,  but  dead  and  dry  organisms,  in  most  cases,  offer  only  a  most 
dubious  and  xuicertaiu  means  of  identification.  Who,  for  example,  would  under- 
take to  determine  what  is  included  under  the  name  Bacterium  iermo  in  von  Thue- 
men's  dried  collection,  No.  1000  ?  The  name  Bacterium  gummis  affords  a  good 
example  of  what  the  writer  has  in  mind.  Bacillus  vasculartim  solani,  Bacillus  cauliv- 
orus,  Bacillus  gossypina,  and  Micrococcus  pellucidus  are  also  examples  of  names  given 
unaccompanied  by  any  proper  description  of  the  organism.  Many  additional  ones 
might  be  cited.  There  is  no  lack.  To  found,  for  example,  a  new  species  of  rabbit 
on  the  observation  that  a  small  jumping  animal  about  the  size  and  shape  of  a  rabbit 
had  congregated  in  certain  turnip  fields  and  caused  great  damage  and  apparently 
had  destroyed  no  other  plants  would  only  serve  to  provoke  a  smile  or  to  raise  a 
doubt  as  to  the  author's  mental  condition,  and  yet  descriptions  equally  worthless 
are  not  at  all  uncommon  in  systematic  bacteriology.  The  Micrococcus  pellucidus, 
although  published  quite  recently  and  in  the  Comptes  Rendus  of  the  French 
Academy,  is  not  described  any  better.  "  I  find  it  quite  impossible,"  says  Mr.  Stod- 
dert,  "  to  identify  many  species  from  published  descriptions."  Numerous  complaints 
of  this  sort,  made  in  recent  years  by  well-trained  and  competent  men,  sufficiently 
indicate  the  necessity  of  a  thoroughgoing  reform. 

Various  more  or  less  arbitrary  dates  have  been  assumed  by  zoologists  and 
botanists  as  the  proper  beginning  of  species  priority,  none  of  which  can  be  used  in 
bacteriology.  In  the  opinion  of  the  writer  the  only  proper  starting  point  is  from 
the  time  when  bacteriologists  were  first  able  to  make  and  easily  maintain  pure 
cultures  of  any  given  organism,  namely,  from  the  discovery  of  the  poured-plate 
method  of  isolation  in  the  year  1881.  Nearly  all  species  characterization  prior 
to  this  time  is  a  cloudland  of  uncertainty,  and  while  it  may  be  possible  fifty 
or  a  hundred  years  from  now,  when  the  whole  field  of  bacteriology,  as  we  now 
understand  it,  shall  have  been  thoroughly  worked  over,  to  decide  by  the  doctrine  of 
exclusion,  with  some  degree  of  probability,  what  was  meant  by  certain  old  names, 
nothing  whatever  can  to-day  be  made  out  of  the  description  accompanying  these 
names.  And  here  I  wish  to  register  a  protest  against  anything  of  this  nature  ever 
being  done.  If,  in  his  own  generation,  a  name  can  not  be  associated  beyond  doubt 


• 

156  BACTERIA    IN    RELATION   TO    PLANT   DISEASES. 

with  a  particular  organism  by  means  of  an  author's  description  or  figures  or  collected 
specimens,  then  this  name  should  disappear,  never  to  be  revived.  Societies  of 
bacteriologists  should  unite  in  the  near  future  on  some  authoritative  date  for  the 
beginning  of  species  priority,  so  that  some  sort  of  stability  may  be  guaranteed  to 
the  nomenclature  of  the  future. 

In  the  way  of  generic  nomenclature  there  is  not  much  of  value  prior  to  Cohn's 
first  great  paper  in  the  year  1872.  It  seems  perhaps  rather  commonplace  reading 
now,  but  it  really  marked  a  great  advance  and  was  the  result  of  twenty  years  of 
diligent  inquiry.  Inasmuch  as  there  is  no  present  agreement  among  bacteriologists 
as  to  the  limits  of  common  genera,  the  same  genus  name  being  used  with  very 
different  meanings  by  different  writers,  it  appears  worth  while  to  discuss  the  subject 
of  genera  at  some  length. 

At  the  outset  three  principal  inquiries  arise.  First,  what  character  or  congeries 
of  characters  shall  be  considered  of  generic  value ;  second,  what  generic  names  shall 
be  used ;  third,  what  meaning  shall  be  attached  to  these  names  ? 

In  the  description  of  species  it  is  necessary  to  draw  very  largely  upon  physio- 
logical characters,  but  it  will  not  be  disputed,  I  think  (certainly  not  by  naturalists), 
that  genera  ought  to  be  founded,  if  possible,  entirely  upon  morphological  characters, 
in  conformity  with  the  usages  of  other  branches  of  natural  history.  Physiological 
characteristics  may  be  used  to  help  out  our  description  of  sub-generic  groups,  such 
as  the  yellow  Bacterium  (Pseudomonas)  group,  the  green-fluorescent  Bacterium 
(Pseudomotias)  group,  the  hog-cholera  group,  the  hay-bacillus  group,  the  Proteus 
group,  the  Tyrothrix  group,  the  Urobacillus  group,  etc.,  but  morphology  appears  to 
be  sufficient  to  distinguish  the  genera. 

Quite  dissimilar  organisms  are  still  put  by  many  writers  under  the  same  genus 
name,  but  the  tendency  to  carefully  discriminate  is  on  the  increase,  and  before  many 
years,  it  is  safe  to  say,  writers  on  bacteria  will  be  using  generic  names  with  a  definite 
morphological  meaning.  Certain  it  is  that  we  can  not  go  on  much  longer  calling 
any  rod-shaped  organism  Bacillus  or  Bacterium  interchangeably,  or  putting  it  into 
the  one  genus  or  the  other  according  as  we  happen  or  do  not  happen  to  find  it 
producing  endospores,  or  growing  as  a  short  rod  or  as  a  filament.  Some  light  may 
come  from  considering  with  what  meaning  such  generic  terms  were  originally  used. 
Matters  would  also  be  much  simplified  by  accepting  1872,  the  date  of  the  appear- 
ance of  Cohn's  first  great  paper,  as  the  proper  date  for  the  beginning  of  generic 
nomenclature  of  the  bacteria,  using  only  such  earlier  names  as  he  accepted,  emend- 
ing his  conception  of  these  genera  in  such  ways  as  experience  has  shown  to  be 
necessary,  and  adding  new  names  from  time  to  time  as  new  genera  are  discovered. 
If  some  date  is  not  settled  upon  in  the  near  future,  then  we  may  expect  an  attempt 
to  substitute  certain  names  which  have  not  been  at  all  used  for  the  last  thirty  years, 
i.  e.,  since  bacteriology  became  a  science,  for  those  which  are  now  in  common  use. 
The  confusion  which  would  result  from  an  attempt  of  this  sort  and  the  utter  useless- 
ness  of  making  such  a  change  are  sufficient  grounds  for  desiring  an  authoritative 
expression  of  opinion  on  the  part  of  organized  bodies  of  men  cultivating  this 
branch  of  science  before  we  are  precipitated  into  any  such  confusion.  The  question 
raised  is  this :  Shall  we  abandon  modern  generic  names  given  to  definite,  well-known, 
and  easily  recognizable  organisms  for  old  names  given  before  there  was  any  science 


NOMENCLATURE   AND    CLASSIFICATIONS. 


157 


of  bacteriology,  not  now  in  use,  and  most  of  which  can  not  be  attached  with  certainty 
to  any  specific  organism,  or  to  any  definite  group  of  organisms? 

From  time  to  time,  as  new  discoveries  have  been  made,  our  views  as  to  what 
should  be  considered  generic  characters  have  undergone  decided  modifications,  as 
everyone  knows  who  is  familiar  with  the  various  writings  on  systematic  bacteri- 
ology, especially  those  which  have  been  the  most  widely  read  and  have  exerted  the 
most  influence.  A  full  discussion  of  all  the  various  problems  of  generic  nomencla- 
ture is  not,  however,  contemplated  in  this  connection.  It  is  safe  to  predict  that  no 
system  now  extant  can  be  looked  upon  as  a  finality,  since  we  know  as  yet  too  little 
about  these  numerous  and  variable  organisms  to  devise  an  altogether  consistent 
system.  Classifications  are  conveniences,  nothing  more.  Some  conform  more 
nearly  than  others  to  the  observed  facts,  but  none  are  perfect  or,  from  the  nature  of 
the  case,  can  ever  be  final.  What  the  future  may  have  in  store  no  one  can  tell. 
There  will  undoubtedly  be  many  surprising  discoveries,  and  recent  attempts  at 
classification  may  then  appear  very  crude.  Our  concern,  however,  is  chiefly  with 
the  present  and  with  knowledge  as  it  appears  to-day. 

On  the  whole,  the  classification  of  Migula,  which  was  proposed  in  October,  1894, 
and  is  outlined  at  length  and  applied  to  most  of  the  well-recognized  forms,  in  his 
beautiful  great  work,  "  System  der  Bakterieu,"  appeals  to  me  most  strongly.  Up 
to  this  time  the  writer  has  followed  this  system  in  his  own  publications  and  will 
continue  to  do  so,  with  certain  modifications,  until  some  distinctly  better  system 
makes  its  appearance.  This  system  is  based  on  the  flagella  and  is  much  more 
workable  than  one  based  on  spores,  or  on  a  combination  of  these  two  characters. 
The  presence  or  absence  of  flagella  and  their  position  on  the  body  are  used  by 
Migula  as  generic  characters. 

In  1895  Dr.  Alfred  Fischer  also  propounded  a  new  system  of  classification 
based  on  spores  and  flagella.  This  system  was  republished  in  1897,  with  material 
modifications,  in  his  "  Vorlesungen  iiber  Bakterien,"  and  is  modified  still  further  in 
the  second  edition  of  that  work.  In  the  non-twisted,  rod-shaped  bacteria  use  is 
made  of  the  flagella  to  separate  the  subfamilies,  while  the  generic  characters  are 
derived  from  certain  phenomena  incident  to  spore-formation.  The  following  table 
of  1 7  genera,  taken  from  his  first  paper,  shows  this  system  at  a  glance : 

FISCHER'S  TABELLARISCHE  UEBERSICHT  DER  BACILLACEBN. 


Sub-families. 

Flagella. 

Shape  of  rods  containing  endospores. 

Arthrospores. 

Cylindric. 

Spindle-form. 

Clavate. 

Bacillei  
Bactriniei..  . 

None  

Bacillus  
Bactrinium.. 

Paracloster.  . 
Clostrinium.  . 

Paraplectrum..  . 
Plectrinium  

Arthrobacter. 
Arthrobactrinium. 

One  polar 
flagellum. 

Bactrillei.  .  . 

Polar  fla- 
gella tuft. 

Bactrillum.  .  . 

Clostrillum.  . 

Plectrillum  

Arthrobactrillum. 

Bactridiei..  . 

Diffuse 
flagella. 

Bactridium.  . 

Clostridium.  . 

Plectridium 
Diplectridium. 

Arthrobactridium. 

158  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

Concerning  this  classification  it  should  be  pointed  out  that  large  groups  of  bac- 
teria are  omitted  altogether,  namely,  those  which  produce  neither  endospores  nor 
arthrospores.  This,  so  far  as  we  yet  know,  includes  nearly  all  the  plant  parasites. 
About  one-half  of  the  genera  were  hypothetical  at  the  time  the  paper  was  published, 
i.  e.,  not  founded  on  any  organism,  as  I  have  already  pointed  out  in  another  place.* 
The  question  of  whether  an  endospore-bearing  rod  is  or  is  not  swollen  around  the 
spore  is  often  difficult  to  determine,  and  as  Migula  and  L/ehmann  &  Neumann  have 
pointed  out,  the  endospore-bearing  rods  in  some  species  may  be  either  cylindric  or 
spindle-form,  or  bear  the  spores  in  the  middle  or  at  one  end.  The  whole  question  of 
the  existence  of  arthrospores  is  still  a  matter  of  doubt.  Closely-related  forms,  and 
even  the  same  species,  may  possess  one  or  several  polar  flagella.  The  genus  Bacillus 
was  founded  by  Colin  on  Bacillus  subtilis,  which  is  now  known  to  have  peritrichiate 
flagella,  Bacillus  ulna,  also  actively  motile,  and  B.  anthracis,  which  is  non-motile. 
Inasmuch  as  Cohn's  studies  were  made  chiefly  on  B.  subtilis,  he  having  never 
studied  B.  anthracis,  but  only  including  it  as  a  sort  of  afterthought,  for  the  sake  of 
completeness,  and  because  Bacillus  subtilis  is  the  first  one  described,  it  seems  only 
proper  that  the  term  Bacillus  should  be  restricted  to  motile  forms  resembling  the 
hay  bacillus,  z.  e.,  those  having  diffuse  flagella,  and  should  not  be  transferred  to  the 
non-motile  forms.  For  the  hay  bacillus  and  similar  forms  Dr.  Fischer  has  used  the 
name  Bactridium.  This  name,  however,  is  inadmissible  because  preoccupied  and 
that,  too,  whether  bacteria  be  considered  as  plants  or  animals.  Bactridium  has  been 
used  as  a  genus  name  seven  times,  as  follows  : 

Bactridium  Kunze,  1817:  For  fungi,  n  species  of  which  are  recognized  in 
Saccardo's  Sylloge  Fungorum. 

Bactridium  Salisb.,  used  in  1839  as  a  sectional  name  under  Erica  (DC.  Prod.), 
and  also  in  1889  by  Drude  in  Engler  &  Prantl's  Die  Natiir.  Pflanzenfamilien.  It 
is  said  by  Baillon  to  be  a  synonym  of  Syringodea  Benth.  Bentham  reduced  Don's 
genus  Syringodea  to  a  section  of  Erica. 

Bactridium  LeConte,  1861:  Col.,  p.  86  MS.;  Bactridium  Sauss.,i863,  Orthop.  M. 
Scudder  :  Genera  in  Zoology. 

Bacteridium  Davaine,  1868  :    For  the  organism  causing  anthrax. 

Bactridium  Schroeter,  1872  :  For  Micrococcus  prodigiosus  and  various  other 
pigment-bearing  bacteria,  most  of  which  have  since  been  included  under  Bacillus. 

Bactridium  Fischer,  1895  :  For  Bacillus  subtilis,  B.  megaterium,  B.  typhosus, 
etc. ;  B.  typhosus,  however,  being  non-sporiferous,  so  far  as  known,  has  logically  no 
place  in  Fischer's  original  classification,  as  already  pointed  out,  since  the  mere  fact 
of  the  absence  of  endospores  does  not  presuppose  the  existence  of  arthrospores. 
The  same  remark  applies  to  Bacillus  amylovorus  and  to  many  other  species. 

Dr.  Fischer  himself  knew  of  the  existence  of  Kunze's  genus  Bactridium,  and 
refers  to  it,  but  he  does  not  appear  to  have  known  of  Davaine's  use  of  the  word  for 
the  anthrax  organism.  He  thinks  that  Kunze's  "rare,  little-known  fungi"  are  so 
different  that  there  will  be  no  confusion,  and  insists  on  using  the  word  with  an 
entirely  different  meaning  for  the  most  curious  of  all  reasons,  viz,  "um  die  Harmonic 

*Review  in  Am.  Naturalist. 


NOMENCLATURE   AND    CLASSIFICATIONS.  159 

der  Nomenclature  nicht  zu  storen."  The  only  possible  ground  on  which  such  use 
could  be  defended  is  that  bacteria  are  so  different  from  plants  and  animals  that  dupli- 
cation of  generic  names  is  not  a  matter  of  any  consequence.  If  this  were  so,  then 
he  should,  nevertheless,  according  to  all  recognized  rules  of  nomenclature,  have  used 
the  word  with  a  different  concept,  since  the  first  use  of  Bacteridium  in  this  group 
was  to  designate  a  non-motile  organism.  Davaine  used  this  word  long  ago  in  a 
perfectly  plain  and  legitimate  way  to  separate  the  non-motile  from  the  motile  forms, 
and  the  organism,  which  he  studied  most  carefully  as  the  type  of  his  new  genus, 
is  that  which  was  only  some  years  afterwards  designated  Bacillus  anthracis,  namely, 
in  1872,  when  Cohn  formed  his  new  genus  Bacillus.  If  the  genus  Bacteridium  is 
to  stand  at  all  as  a  bacterial  genus,  it  must  be  used  for  Bacillus  anthracis  and  its  con- 
geners, and  it  is  an  eminently  proper  name  for  these  organisms,  provided  bacteri- 
ologists can  bring  themselves  to  think,  with  Fischer,  that  this  name  is  not  too 
close  to  the  fungous  genus  Bactridmm  Kunze  on  the  one  hand  or  to  the  animal 
genera  on  the  other,  which  have  priority.  The  writer  does  not  share  this  opinion. 
There  would  seem  to  be  also  an  older  name  than  Dr.  Fischer's  Plectrinium  for 
monotrichiate  bacteria  having  the  spore  borne  in  a  swollen  end,  viz,  Trdcul's 
Urocephaliim.  This,  according  to  Trecul,  was  a  motile  bacterium  coloring  deep 
blue  with  iodine,  Trdcul  describes  it  as  0.02  mm.  long,  "a  queue  flexueuse,"  with 
distinct  spore  formation  at  one  end  which  was  enlarged.  The  subfamily  name 
Bactridei  is  also  open  to  objection  because  preoccupied.  There  is  a  genus  of  palms, 
Bactris,  and  in  1889  Drude,  in  "Die  Natiirlichen  Pflanzenfamilien,"  used  the  sub- 
tribal  name  Bactrideae,  which  is  the  same  word  as  Bactridei. 

Dr.  Fischer  has  not  helped  matters  by  the  modifications  introduced  into  his 
"  Vorlesungen "  as  the  result  of  criticism,  since  he  has  destroyed  the  logical  con- 
sistency of  his  system  by  including  sporiferous  and  non-sporiferous  forms  under  the 
same  genus  name. 

MIGULA'S  CLASSIFICATION  OF  THE  BACTERIA. 

The  bacteria  are  phycochrom-free  schizomycetous  plants  with  division  in  one, 
two,  or  three  directions  of  space  ;  reproduction  by  vegetative  multiplication.  Resting 
stages  in  the  form  of  endospores  are  produced  in  many  sorts.  Motility  due  to 
flagella  occurs  in  some  genera.  In  Beggiatoa  and  Spirochaeta  the  organs  of  motion 
are  unknown. 

I.    Order  EUBACTERIA. 

Cells  without  any  ' '  Centralkorper ' '  and  free  from  sulphur  and  bacteriopurpurin  ; 
colorless  or  faintly  colored,  also  chlorophyll-green. 

1.    Family  COCCACEAE  Zopf  emend.  Mig. 

Cells  globose  in  a  free  state  ;  in  stages  of  division  often  somewhat  elliptical. 
Division  in  one,  two,  or  three  directions  of  space  without  previous  elongation  of  cell. 
If  the  cells  remain  united  after  division,  they  are  frequently  flattened  at  points  of 
cohesion.  In  all  Coccaceae  with  cells  large  enough  for  observation  septation  takes  place 
in  the  globose  state  before  there  is  any  elongation  perpendicular  to  plane  of  division. 
Only  a  few  species  are  motile. 


l6o  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

Streptococcus  Billroth. 

Cells  globose  and  without  organs  of  motion.     Division  in  only  one  direction  of 
space.     When  the  cells  remain  united  after  division,  moniliform  chains  are  produced, 
or  diplococcus  forms,  but  the  latter  also  occur  in  other  genera  of  Coccaceae.    Doubtful 
if  any  spore  formation. 
Micrococcus  (Hallier)  Cohn. 

Cells  globose  in  a  free  state.  Division  in  two  directions  of  space  without  previous 
elongation  of  cell.  No  organs  of  motion.  Endospore  formation  not  positively  demon- 
strated and  probably  wanting.  When  the  cells  remain  together  after  division  Meris- 
mopaedia-like  plates  may  be  formed,  in  which  case  the  contiguous  cell- walls  may  be 
flattened. 
Sarcina  Goodsir. 

Cells  globose,  in  a  free  state.     Division  in  three  directions  perpendicular  to  each 
other.     No  organs  of  motion.     Spore  formation  doubtful.     If  the  cells  remain  united 
after  division,  bale-like  constricted  packetsare  formed  ;  frequently  these  do  not  appear, 
as  the  nutrient  medium  has  the  greatest  influence  on  the  form  of  the  cell-unions. 
Planococcus  Migula. 

Cells  globose  but  usually  adhering  in  twos  or  fours  with  points  of  contact  flat- 
tened.   Division  in  two  directions  of  space,  as  in  Micrococcus.      Motile  by  means  of 
one  or  two  long,  wavy-bent  flagella.     Spore  formation  unknown. 
Planosarcina  Migula. 

Free  cells  globose.  Division  in  three  directions  of  space,  as  in  Sarcina.  Motile 
by  means  of  long  or  short  flagella.  Apparently  only  one  flagellum  to  each  cell.  No 
endospores.  Usually  the  cells  remain  united  after  division  as  diplococci  or  tetracocci, 
but  seldom  in  the  form  of  packets. 

2.     Family   BACTERIACEAE. 

Cells  longer  or  shorter  cylindric,  straight,  or  at  least  never  spirally  twisted. 
Division  in  one  direction  only,  viz,  perpendicular  to  long  axis,  and  only  after  a  pre- 
liminary elongation  of  the  rod.  In  some  species  the  rods  separate  early ;  in  others 
they  remain  united  for  a  considerable  time  as  longer  or  shorter  threads.  A  single  cell, 
so  far  as  known ,  does  not  immediately  break  up  into  two  daughter  cells  when  the 
first  septum  is  formed,  but  remains  as  a  single  rod  until  additional  septa  are  laid  down. 
In  some  species  the  cells  may  be  very  short,  so  as  to  superficially  resemble  Strepto- 
cocci, but  an  exact  study  of  the  cell-division  enables  one  to  distinguish  with  certainty. 
Bacterium  Ehrenberg  (char,  emend.). 

Cells  cylindric,  longer  or  shorter,  often  forming  threads  of  considerable  length. 
Without  organs  of  motion.  Endospore  formation  occurs  in  many  species,  but  appears 
to  be  entirely  wanting  in  others.  In  many  they  may  yet  be  discovered  when  the 
organisms  are  exposed  to  suitable  environments. 

Bacillus  Cohn  (char,  emend.). 

Cells  straight,  rod-shaped  to  ovoid,  longer  or  shorter,  sometimes  united  into  quite 
long  threads.  Motile  by  means  of  wavy-bent  flagella  which  are  scattered  over  the  whole 
body.  Endospore  formation  frequent.  In  most  species  motility  occurs  only  during  a 
definite  period  of  development,  which  is  very  brief  in  some  species  and  very  long 
in  others. 
Pseudomonas  Migula. 

Cells  cylindric,  shorter  or  longer,  sometimes  forming  threads.  Motile  by  means 
of  polar  flagella.  The  number  of  flagella  on  a  pole  varies  from  i  to  10  ;  most  frequently 
it  is  i,  or  3  to  6.  Endospore  formation  certainly  occurs  in  some  sorts,  but  is  rare. 


NOMENCLATURE   AND    CLASSIFICATIONS.  l6l 

A  separation  into  two  genera  does  not  appear  to  be  desirable  at  the  present  time. 
No  such  difference  exists  inside  this  genus  as  there  is  between  the  genus  Microspira, 
which  only  exceptionally  has  more  than  one  flagellum  on  the  pole,  and  the  genus 
Spirillum,  which  has  many  polar  flagella.  Between  the  one-flagellate  and  many- 
flagellate  forms  there  are  all  sorts  of  transitions  in  the  genus  Pseudomonas.  Possibly 
the  boundary  between  Pseudomonas  and  Microspira  is  artificial.  Slight  crooking  of 
the  rods,  especially  in  stained  preparations,  has  been  observed  in  many  species  of 
Pseudomonas,  and  it  is  not  always  possible  to  decide  whether  a  one-flagellate  form 
belongs  to  this  genus  or  to  Microspira.  Ordinarily,  a  decision  may  be  reached  by 
observing  the  shape  of  the  threads  or  chains,  those  of  Pseudomonas  never  being 
twisted  into  the  form  of  a  screw. 

3.    Family  SPIRILLACEAE  (Screwbacteria) . 

Cells  spirally  wound  or  representing  a  portion  of  the  turn  of  a  spiral,  in  which 
latter  case  the  entire  spiral  is  visible  only  when  several  cells  remain  united.  Endo- 
spore  formation  established  for  some  species,  but  rare.  Apparently,  in  some  of  the 
larger  species  resting  forms  are  also  produced  by  the  breaking  up  of  the  rods  into 
short  segments  which  envelop  themselves  in  a  gelatinous  membrane.  Usually  motile. 
Division  only  in  one  direction  of  space  perpendicular  to  the  long  axis. 

Spirosoma  Migula. 

Cells  generally  rather  large,  spirally  bent,  rigid,  and  without  organs  of  motion. 
Cells  single,  free,  or  united  into  small  gelatinous  families. 

(1)  Subgenus  EUSPIROSOMA  :    Cells  single  or  united  into  a  spirally  twisted  thread. 
Free,  that  is,  not  inclosed  in  any  gelatinous  envelope. 

(2)  Subgenus  MYCONOSTOC:    Cells  single  or  united  into  spiral  threads,  which  are 
surrounded  by  a  roundish  general  envelope. 

Microspira  Schroter. 

Cells  bent  like  a  comma  or  sausage,  rigid,  single  or  several  united,  in  which  latter 
case  screws  or  S-shaped  figures  are  produced.  Motile  by  means  of  i  wavy-bent  polar 
flagellum  (rarely  2  or  3  flagella).  The  flagellum  is  usually  not  much  longer  than  the 
cell.  Endospore  formation  has  thus  far  not  been  demonstrated.  Many  authors  do 
not  distinguish  between  this  genus  and  the  next. 

Spirillum  Ehrenberg. 

Cells  rigid,  rods  of  various  thicknesses,  length,  and  pitch  of  the  spiral,  forming 
either  long  screws  or  only  portions  of  a  turn.  Cells  motile  by  means  of  a  tuft  of  polar 
flagella  (5  to  20),  which  are  mostly  half  circular,  rarely  wavy-bent.  These  flagella 
occur  on  one  or  both  poles.  Their  number  varies  greatly  and  is  difficult  to  determine, 
since  in  stained  preparations  several  are  often  united  into  a  common  strand.  Endo- 
spore formation  has  been  observed  in  some  species.  There  are  many  undescribed  species. 

Spirochaeta  Ehrenberg. 

Cells  thin,  mostly  quite  long,  motile  and  flexible,  winding  snake-like,  but  also 
moving  in  the  manner  of  a  screw.  Organs  of  motion  unknown.  Endospore  forma- 
tion not  observed. 

Nearly  related  to  the  Algal  genus  Spirulina,  but  colorless  and  not  segmented  into 
single  cells. 

4.    Family  CHLAMYDOBACTERIACEAE. 

Cells  cylindric,  united  into  threads  which  are  surrounded  by  a  sheath.  Repro- 
duction by  means  of  motile  or  non-motile  conidia  which  arise  directly  from  the  vege- 
tative cells,  and  without  passing  through  any  resting  stage  grow  into  new  threads. 


l62  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

Chlamydothrix  Migula. 

Cells  cylindric,  non-motile,  arranged  in  unbranched  threads,  with  a  sheath  of 
varying  thickness.  Frequently  the  septation  of  the  threads  is  only  demonstrable  after 
the  use  of  reagents.  Reproduction  by  means  of  non-motile,  roundish  or  ovoid 
conidia,  which  arise  directly  from  the  vegetative  cells.  Syn . :  Streptothrix  (Cohn)  Mig. , 
Leptothrix  Kiitz.  exp.',  and  Gallionella  Ehrenberg  exp. 
Crenothrix  Cohn. 

Cells  united  into  unbranched  filaments,  attached,  and  gradually  enlarging  toward 
the  free  end,  i.  e.,  with  a  distinction  between  base  and  apex.  Sheath  rather  thick. 
In  iron  waters  the  old  and  empty  sheaths  are  permeated  by  ironoxidhydrate.  Cells 
cylindric  or  flat  discoidal.  Multiplication  by  non-motile  (mostly  roundish)  conidia, 
which  arise  from  the  vegetative  cells  by  division  and  rounding  off.  For  this  purpose 
the  cells  of  the  thicker  threads  divide  in  three  directions  of  space,  those  of  the  thinner 
threads  only  perpendicularly  to  the  long  axis  of  the  thread.  The  conidia  are  discharged 
and  germinate  immediately,  often  on  the  sheath  of  the  mother-thread.  Only  one 
species  known. 
Phragmidiothrix  Engler. 

Cells  cylindric,  later  discoidal,  forming  threads  100  ^  long  and  3  to  12  ju  thick, 
with  a  very  delicate,  scarcely  visible  sheath.  Multiplication  by  non-motile  conidia, 
which  arise  from  the  vegetative  cell  by  division  in  three  directions  of  space  and 
rounding  off.  Perhaps  to  be  united  with  Crenothrix,  if  the  projections  observed  by 
Engler  are  not  branches  but  epiphytes.  Only  one  sort  known. 
Sphaerotilus  Kutzing  (1833). 

Cells  cylindric,  enveloped  in  sheaths,  forming  dichotomously  branched  threads 
with  no  differentiation  into  base  and  apex.  Multiplication  by  means  of  conidia, 
which  swarm  out  of  the  sheath,  attach  themselves  anywhere,  and  immediately  grow 
out  into  new  threads.  The  conidia  possess  a  tuft  of  flagella  inserted  sidewise  under 
one  pole.* 

Genera,  the  systematic  position  of  which  is  doubtful :  Spiromonas  Perty  ;  Spiro- 
discus  Ehrenberg ;  Achromatium  Schewiakoff ;  Newskia  Famintzin ;  Streblothrichia 
Guignard. 

II.    Order  THIOBACTERIA. 

Cells  without  any  "Centralkorper,"  but  with  sulphur  inclusions.  Colorless,  or 
pigmented  rose,  red,  or  violet  by  bacteriopurpurin ;  never  green. 

1.    Family  BEGGIATOACEAE. 

Filamentous  bacteria,  destitute  of  bacteriopurpurin.  Division  of  cells  in  one 
direction  of  space,  viz,  perpendicular  to  the  long  axis. 

Thiothrix  Winogradsky. 

Threads  attached,  not  uniformly  thick,  enveloped  in  a  delicate  sheath  which  is 
not  easily  demonstrable,  non-motile,  contents  containing  sulphur  granules.  The 
threads  produce  rod-shaped  conidia  at  their  end.  These  conidia,  which  are  self-motile 
by  means  of  a  slow,  creeping  motion,  attach  themselves  by  one  end  to  any  sort  of 
substratum,  extrude  a  slime-cushion  at  the  base,  bend  over  ordinarily  in  their  middle 
to  a  nearly  right  angle  and  grow  into  a  new  thread.  Habitat,  hot  sulphur  springs. 
Beggiatoa  Trevisan. 

Threads  destitute  of  a  sheath,  formed  of  flat  discoidal  cells,  free,  i.  e.,  not  attached. 
Multiplication  by  folding  and  separation  of  the  threads.  Motile  by  means  of  an 

'Streptothrix  and  Cladothrix  are  omitted  from  the  second  volume,  the  species,  so  far  as  they  repre- 
sent bacteria,  being  distributed  in  other  genera.  C.  dichotoma  becomes  SphcKrotilus  dichotomus. 


NOMENCLATURE    AND    CLASSIFICATIONS.  163 

undulating  membrane,  as  in  Oscillaria.  The  organism  creeps  along,  but  at  the  same 
time  rotates  around  the  long  axis,  mostly  with  a  swinging  of  one  or  both  free  ends. 
Habitat,  hot  sulphur  springs  and  other  fluids  in  which  hydrogen  sulphide  is  developed. 
No  reliable  method  is  yet  known  for  the  separation  of  the  species.  The  number  and 
size  of  the  included  sulphur-granules  are  not  of  specific  value.  They  depend  on  the 
amount  of  hydrogen  sulphide  in  the  water. 

2.   Family  RHODOBACTERIACEAE. 

Cell-contents  rose,  red,  or  violet,  from  the  presence  of  bacteriopurpurin .  Sulphur 
granules  are  also  included. 

Classification  still  very  artificial,  owing  to  imperfect  knowledge.  Author  follows 
Winogradsky. 

(I)  Subfamily  THIOCAPSACEAE. 

Cells  united  into  families.     Division  of  the  cells  in  three  directions  of  space. 
Thiocystis  Winogradsky. 

Families  small,  compact,  enveloped  singly  or  several  together  in  a  gelatinous 
cyst,  capable  of  swarming.  When  the  families  have  reached  a  definite  size  they 
escape  from  the  gelatinous  cyst,  the  latter  either  swelling  and  softening  uniformly  or 
at  some  particular  spot.  The  escaped  cells  either  pass  iuto  the  swarm  stage  or  unite 
into  a  larger  fused  complex  of  families,  the  individual  cells  of  which  separate  and 
swim  away  only  after  a  long  time,  and  by  means  of  much  vigorous  struggling. 

Thiocapsa  Winogradsky. 

Non-swarming,  globose  cells  spread  out  upon  the  substratum  in  flat  families, 
which  are  loosely  enveloped  in  a  common  gelatin.     The   membrane  is  split  by  the 
growth  of  the  family,  and  the  cells  are  separated  as  if  by  the  swelling  of  an  inter- 
mediate substance. 
Thiosarcina  Winogradsky. 

Non-swarming  cells  arranged  in  packet-shaped  families,  corresponding  to  the 
genus  Sarcina  in  the  Eubacteriaceae. 

(II)  Subfamily  LAMPROCYSTACEAE. 

Cells  united  into  families.     Divisions  of  the  cells,  first  in  three  and  then  in  two 
directions  of  space. 
Lamprocystis  Schroter. 

Families  at  first  solid,  then  globose-hollow,  becoming  perforated  net-form;  cells 
finally  separating  into  small  groups  which  are  capable  of  swarming. 

(Ill)  Subfamily  THIOPEDIACEAE. 

Cells  united  into  families.     Divisions  in  two  directions  of  space. 
Thiopedia  Winogradsky. 

Families  tabular,  formed  of  cells  arranged  in  fours  and  capable  of  swarming. 

(IV)    Subfamily  AMOEBOBACTERIACEAE. 

Cells  united  into  families.     Division  in  one  direction  of  space. 
Amoebobacter  Winogradsky. 

Cells  connected  by  plasma  threads.  Families  amoeboid  motile.  The  cell  families 
slowly  change  form,  the  cells  drawing  together  into  a  heap  or  spreading  out  widely, 
thus  bringing  about  a  change  in  the  shape  of  the  whole  family.  In  a  resting  condi- 
tion a  common  gelatin  is  extruded,  the  surface  of  which  becomes  a  firm  membrane. 


164  BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 

Thiothece  Winogradsky. 

Families  inclosed  in  a  thick  gelatinous  cyst.     Cells  capable  of  swarming  and 
very  loosely  embedded  in  a  common  gelatin.    When  the  swarm  stage  supervenes,  the 
cells  lie  more  loosely,  the  gelatin  is  swollen,  and  the  cells  swarm  out  singly  and 
rather  irregularly. 
Thiodictyon  Winogradsky. 

Families  consisting  of  rod-shaped  cells  having  their  ends  united  into  a  net.  Out 
of  an  originally  compact  mass  of  rods  there  gradually  results  from  rearrangement  a 
Hydrodictyon-like  cell-union,  which,  under  unfavorable  conditions,  may  again  draw 
together  into  a  compact  mass  of  rods.  The  multiplication  of  the  families  results  from 
division  or  by  loosening  of  slowly  motile  small  cell-colonies. 
Thiopolycoccus  Winogradsky. 

Families  solid,  non-motile,  consisting  of  small  cells  closely  pressed  together. 
Multiplication  of  the  colonies  by  the  breaking  up  of  the  surface  into  numerous  short 
shreds  and  lobes  which  continue  to  split  up  into  smaller  heaps. 

(V)    Subfamily  CHROMATIACEAE. 

Cells  free,  capable  of  swarming  at  any  time. 
Chromatium  Perty. 

Cells  moderately  thick,  cylindric-elliptical  or  elliptical.     Polar  flagella. 
Rhabdochromatium  Winogradsky. 

Cells  rod-shaped  or  spindle-form,  with  flagella  on  the  poles. 
Thiospirillum. 

Cells  spirally  twisted. 

To  the  preceding  may  be  added  a  third  order,  Myxobacteriacese,  which  is  not 
discussed  critically  by  Migula  and  is  not  adequately  described  in  any  of  the  text- 
books. The  following  general  characters  are  taken  from  the  papers  of  Dr.  Roland 
Thaxter,  to  whom  we  owe  our  knowledge  of  these  curious  and  interesting  organ- 
isms. The  most  recent  paper  is  by  Erwin  Baur,  Myxobakterien-Studien,  Archiv  fur 
Protistenkunde,  V  Bd.,  I  Heft,  pp.  92-121,  i  pi. 

MYXOBACTERIACEAE. 

Motile,  rod-like  organisms,  multiplying  by  fission,  secreting  a  gelatinous  base, 
and  forming  pseudoplasmodium-like  aggregations  before  passing-  into  a  more  or  less 
highly  developed  cyst-producing,  resting  state,  in  which  the  rods  may  become  encysted 
in  groups  without  modification,  or  may  be  converted  into  spore-masses.  The  vegeta- 
tive rods,  which  vary  little  in  size  and  form  in  the  different  genera  and  species,  are 
typically  elongate,  sometimes  reaching  15  /<  in  length.  Cell-division  follows  an 
elongation  and  nearly  medium  constriction  of  the  rods,  which,  except  at  the  moment 
of  division,  are  always  separate;  never  united  in  chains.  A  slow,  sliding  locomotion 
and  a  Beggiatoa-like,  bending  motion  is  characteristic  of  the  active  rods.  Organs 
of  motion  have  not  been  detected.  In  all  species,  with  one  exception,  the  rods, 
when  seen  in  masses,  are  more  or  less  distinctly  reddish.  A  distinct,  firm,  hyaline, 
gelatinous  base  is  secreted  by  the  colony  as  it  extends  itself,  over  which  the  individuals 
may  move  or  in  which  they  become  embedded. 

The  vegetative  period,  in  artificial  cultures,  usually  lasts  about  a  week,  or  even 
two  weeks,  but  in  nature  the  production  of  cysts  must  be  more  rapid.  Common  in 
moist  situations  on  dung,  decaying  wood,  fungi,  lichens,  etc.  According  to  Bauer 
they  grow  best  at  30°  C. 


NOMENCLATURE   AND    CLASSIFICATIONS.  165 

In  forms  like  Myxococcus,  in  which  the  rods  are  somewhat  scattered,  the  first 
preparation  for  spore-production  as  seen  under  the  microscope  consists  in  the  appear- 
ance of  groups  of  rods  moving  with  a  circular  tendency,  in  which  the  more  central 
individuals  soon  become  converted  into  spores.  The  formation  of  a  cystophore,  when 
it  occurs,  results  from  the  basal  constriction  of  a  papillate  mass  of  rods  which  pro- 
jects from  the  surface  of  the  colony.  In  the  encysted  condition  there  are  two  classes — 
one  in  which  the  individuals  thus  encysted  show  little  or  no  modification  from  the 
rod-like  vegetative  state,  the  other  in  which  they  are  converted  into  definite  spores. 
They,  however,  seem  to  run  into  one  another. 

The  species  have  been  arranged  under  three  genera,  as  follows  : 
Chondromyces  B.  &  C. 

Rods  forming  free  cysts,  in  which  they  remain  unmodified.    Cysts  various,  sessile, 
or  borne  on  a  more  or  less  highly  developed  cystophore. 
Polyangium  Lk.  (Myxobacter  and  Cystobacter  syn.). 

Rods  forming  large,  rounded  cysts,  one  or  more  free  within  a  gelatinous  matrix 
raised  above  the  substratum. 
Myxococcus  Thaxter, 

Rods  slender,  swarming  together  after  a  vegetative  period  to  form  definite,  more 
or  less  encysted,  sessile  or  stalked  masses  of  coccus-like  spores. 

That  which  appears  least  defensible  in  Migula's  classification  is  his  use  of  the 
word  Bacterium  for  the  anthrax  organism  and  similar  non-motile  bacteria.  If  this 
generic  name  is  to  be  retained,  it  should  be  used  somewhat  as  Ehrenberg  used  it, 
viz,  for  motile  organisms,  and  should  not  be  given  to  entirely  different  non-motile 
forms.  We  have  the  right  to  set  aside  so  much  of  Ehrenberg's  description  as  does 
not  correspond  to  facts,  but  not  more.  We  do  not  know  exactly  what  Ehrenberg 
had  in  mind,  it  is  true,  but  it  certainly  was  not  non-motile  forms  of  the  type  of  the 
anthrax  organism. 

Provided  one  goes  back  of  Cohn's  time  (1872),  which  the  writer  is  not  disposed 
to  do  in  case  of  the  bacteria,  the  one  species  by  which  the  generic  name  Bacterium 
must  stand  or  fall  is  Bacterium  triloculare  Ehrenberg.  In  size  and  shape,  as  described 
and  figured,  it  agrees  very  well  with  some  of  the  larger  species  of  Pseudomonas 
Migula.  If  we  can  trust  Ehrenberg's  distinct  statements  and  his  plain  figure,  it 
was  provided,  like  most  species  of  Pseudomonas,  with  one  polar  flagellum.  Ehren- 
berg also  figures  and  describes  it  as  trilocular  or  triarticulate.  He  may  have  been 
wrong  in  including  it  among  his  Vibrionides  and  in  figuring  and  describing  it  as 
possessed  of  a  polar  flagellum,  an  organ  difficult  to  make  out  in  unstained  material 
and  with  the  crude  microscopes  in  use  in  his  day,  but,  while  we  bear  this  in  mind, 
we  must  not  forget  that  the  person  who  was  using  these  microscopes  was  no 
ordinary  observer,  but  a  man  with  remarkable  eyesight  and  with  a  genius  for 
observations  of  this  kind.  Moreover,  the  tri-locularity  which  he  observed  may 
have  been  simply  the  organism  in  early  stages  of  division,  which  is  the  more  likely, 
since  he  states  that  he  saw  it  divide,  and  because  in  his  specimens  from  Berlin  he 
sometimes  observed  four  septa  and  sometimes  only  two.  Ehrenberg's  description 
of  the  genus  Bacterium,  taken  as  a  whole,  is  of  course  worthless  for  purposes  of 
modern  classification,  our  ideas  of  generic  values  being  entirely  different  from  his. 
A  few  things  only  come  out  of  the  rubbish  heap  of  this  early  writing  in  a  service- 


1 66  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

able  condition.  The  organism  was  a  short  rod,  multiplying  by  cross-septation, 
possibly  also  by  means  of  spores,  colorless,  sluggishly  motile,  moving,  it  is  said,  by 
means  of  one  polar  flagellum,  and  occurring  abundantly  in  water  containing  rotten 
vegetation,  where  bacteria  would  be  likely  to  abound.  It  should  also  be  noted  that 
it  did  not  take  up  colored  particles,  such  as  indigo  or  carmine,  when  placed  in  water 
containing  these  substances.  If  the  word  Bacterium  is  used,  it  should  be  in  con- 
formity to  these  facts,  or  supposed  facts,  and  may  be  so  used,  I  think,  until  they  are 
shown  to  be  erroneous. 

The  matter  is  simplified,  however,  if  we  start  with  Cohn's  use  of  the  word 
Bacterium  in  the  year  1872.  The  Bacterium  of  Cohn  is  a  certain  Bacterium 
termo.  While  we  are  not  able  to  tie  down  Cohn's  use  of  this  name  to  a  particular 
species,  it  appears  that  we  can  do  it  quite  definitely  to  a  group  of  morphologically 
similar  species.  Much  discredit  has  been  thrown  on  Bacterium  termo  in  modern 
times,  and  it  has  been  left  out  of  many  classifications.  However,  if  one  examines 
into  the  matter,  there  is  no  reasonable  doubt  as  to  what  Cohn  had  in  mind.  His 
Bacterium  termo  was  a  small  schizomycetous  organism  capable  of  growing  freely 
in  Cohn's  nutrient  solution,  containing  acid  potassium  phosphate  and  ammonium 
tartrate.  It  produced  therein  short  rods  (single,  in  pairs,  or  fours  joined  end  to  end) 
and  roundish-lobed  (kugelige-traubige)  white  zoogloese,  together  with  a  greenish 
I  fluorescence.  This  is  Cohn's  statement  and  de  Bary's.  It  did  not 

appear  in  boiled  fluids,  i.  <?.,  was  destitute  of  endospores  (Cohn), 
and  the  motile  rods  were  killed  by  a  short  exposure  to  58°  C. 
(Schroeter).  In  other  words,  it  was  a  non-sporiferous  green- 
fluorescent  organism  possessed  of  a  single  polar  flagellum,  or,  in 
some  cases  perhaps,  provided  with  paired  or  triple  polar  flagella. 
If  we  start  with  Cohn's  classification  in  the  year  1872,  we  may 
p-  |37*  keep  the  name  Bacterium  for  schizomycetous  organisms  of  this 

»  type,  and  at  the  same  time  we  shall  not  be  doing  any  violence  to 

the  older  use  of  the  word  by  Ehrenberg,  who  figures  and  describes  this  kind  of  an 
organism.  This  the  writer  proposes  to  do,  substituting  Bacterium  (Cohn  emend.) 
for  Pseudomonas  Migula  and  for  more  recent  names  proposed  by  others.  Cohn's 
description,  be  it  understood,  is  worthless  for  the  most  part,  but  his  name  Bacterium 
(B.  termo)  is  usable  because  it  can  be  attached  to  a  definite  kind  of  organism.  To  show 
that  Cohn's  use  of  this  word  and  the  writer's  use  of  it  do  not  conflict  with  former 
usage,  Ehrenberg's  descriptions  and  figures  of  Bacterium  are  here  reproduced  from 
the  expensive  and  not  readily  accessible  publications  in  which  they  appeared. 

The  organism  described  as  Bacterium  triloculare  by  Ehrenberg  is  shown  in 
figs.  137  and  138,  and  Ehrenberg's  account  is  summarized  as  follows: 

The  genus  Bacterium  was  founded  by  C.  G.  Ehrenberg  in  1828,  and  was 
characterized  by  him  in  the  Symbolae  Physicae,  Animalia  evertebrata.  The  book  in 
which  this  description  occurs  is  an  unpaged  folio.  On  the  second  page  of  the  text 
proper,  in  a  list  of  species  found  "In  Oasi  lovis  Hammonis  Siwae"  this  genus 

*FiG.  137. — Bacterium  triloculare.     From  Ehrenberg's  Symbolae  Physioe.      Animalia  evertebrata. 
Decas  prima.    Berlin,  1828.     Plate  II,  fig.  6. 


NOMENCLATURE   AND    CLASSIFICATIONS.  167 

is  first  mentioned  as  follows :  "4.  Bacterium  triloculare  n.  g."  The  second  mention 
of  the  name  is  on  the  fourth  page  in  a  list  of  "  Genera  et  Species,  in  Europe  a  me 
nondum  visa."  The  genus  is  described  on  the  sixth  page  as  follows: 

BACTERIUM,  Novum  Genus,  Familia  Vibrioniorum.  Character  Generis  :  Corpus 
polygastricum ?  anenterum?  nudum,  oblongum,  fusiforme  aut  filiforme,  rectum,  mono- 
morphum  (contractione  nunquam  dilatatum),  parum  flexile  (nee  aperte  undatum), 
transverse  in  multas  partes  sponte  dividuum. 

B.  triloculare  nov.  spec.  :  distincte  triloculare  s.  triarticulatum,  subfusiforme, 
hyalinum. 

Phytozoa  Tab.  II.     Libyca  fig.  6. 

Animalculum  i /, '300  lineae  longum,  corpore  tereti.  Articuli  s.  septa  interna 
divisionern  instantem  multiplicem  transversam  indicare  videntur.  Mobile  sed  pigrum 
anirnalculum. 

In  Oasi  lovis  Hammonis  Siwae  observatum,  praeterea  nullibi. 

Bacterii  Generis  physiologia  hucusque  obscura.  Cibo  colorato  ventriculos  replere 
hae  formae  respuunt  ideoque  ad  Polygastrica  non  nisi  dubitanter  et  interim  collocantur. 

Bacterium  simplex  vide  Monas  simplex.  Bacterium  scintillans  vide  Monas  scin- 
tillans. 

In  general,  Ehrenberg's  figures  are  excellent  where  he  could  see  anything  to 
draw,  but  the  seven  figures  of  B.  triloculare  are  very  small  (white  on  a  dark -gray 
background),  and  one  can  make  nothing  out  of  them  beyond  the  fact  that  they  are 
minute  bodies,  3  or  4  times  as  long  as  broad  and  with  two  indistinct  septa.  At  the 
foot  of  the  plate  their  length  is  said  to  be  ^  of  a  "linie."  The  figures  are  more  or 
less  rounded  at  the  ends  and  show  no  constrictions  at  the  septa.  f 

In  1830,  in  his  "Beitrage"  (see  Bibliog.,V),  Ehrenberg  has  the  following  on  the 
genus  Bacterium  : 

Phytozoa.  Classis  I  Polygastrica.  A.  Anentera.  Ordo  I  Nuda.  Family  I  Gymnica. 
Corpore  non  ciliato,  oreciliato  nudove  (p.  37).  Sectio  II.  V!BRIONIA.  Elongata,  in 
se  nunquam  contracta.  Sub-section  c:  Corpore  oblongo.fusiformi  aut  filiformi  (tereti 
aut  triquetro  nee  quadrangulo)  aperte  undatum  non  flexili,  nee  spiral! : 

BACTERIUM,  nov.  Gen. — Haec  genera,  Oscillatories  valde  afYinhi,  ore  nutriri 
nondum  vidi.  n  species  (p.  38). 

In  another  place  the  following  species  are  mentioned,  without  description,  as 
belonging  to  this  new  genus  :  Bacterium  cylindric,  B.  deses,  B.  Enchelys,  B.  fuscum, 
B.  monas,  B.  punctual,  B.  termo,  B.  tremulans.  The  number  1 1  was  apparently 
completed  by  the  three  others,  B.  triloculare,  B.  simplex,  and  B.  scintillans,  not  here 
mentioned.  In  this  paper  Ehrenberg  made  use  of  the  subsequently  oft-quoted  expres- 
sion :  "  Die  Milchstrasse  der  Kleinsten  Organization  geht  durch  die  Gattungen 
Monas,  Vibrio,  Bacterium,  Bodo."  This  idea  appears  to  have  impressed  him  greatly, 
for  he  repeats  it  in  his  "Infusionsthierchen." 

In  1832,  in  Ehrenberg's  "  Geographische  Verbreitung,"  a  paper  which  was  read 
January  10,  1828,  and  written  apparently  before  either  of  the  preceding,  but  not 
printed  until  1832,  three  species  of  Bacterium  are  mentioned,  B.  simplex,  B.trilo- 
culare,  B.  scintillans,  all  as  new  species,  but  there  is  no  description  of  the  genus  or 
reference  to  any  description. 


1 68  BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 

In  1832,  Ehrenberg  again  returns  to  Bacterium  in  a  paper,  "  Ueber  die  Entwicke- 
lung,"  etc.  In  this  paper  six  species  are  included  in  the  genus  Bacterium,  but  four 
of  them  he  now  regarded  as  doubtful : 

Gattung  XII  Bacterium  E.,  Gliederstabchen. 
J)  deutliche  Gliederung. 

1.  B.  articulatum  E.,  Perlenschnurgliederstabchen. 

[Descript.]     Bewegung  zitternd.     Berlin. 

2.  B.  triloculare  H.  und  E.,  dreifachriges  Gliederstabchen. 

JJ)  undeutliche  Gliederung. 

3.  B.  ?  Enchelys  E. 

4.  B.  ?  punctum  E. 

5.  B.  ?  tremulans  E. 

6.  B.  ?  termo  E. 

In  1838,  in  his  magnificent  work,  "  Die  Infusionsthierchen,"  Ehrenberg  gives 
additional  information  respecting  the  genus  Bacterium.  It  is  there  described  as 
follows : 

Die  quergetheilten  gehoren  zu  den  Zitterthierchen  (Vibrionien),  den  langsge- 
theilten  zu  den  Stabthierchen  (Bacillarien)  (p.  2). 
Vierte  Familie:  Zitterthierchen  (Vibrionia,  Vibrionides). 

Character  :  Animalia  filiformia,  distincte  aut  verismiliter  polygastrica,  anentera, 
nuda,  gymnica,  corpore  Monadinorum  uniformi,  divisione  spontanea  imperfecta  (trans- 
versa),  catenatim  consociata,  hinc  filiformia. 

Charactere :  Animaux  filiformes,  distinctement  ou  vraisemblablement  polygas- 
trique,  sans  canal  alimentaire,  sans  carapace,  sans  appendices,  £  corps  uniforme  des 
Monadines,  se  reunissant  par  division  spontanee  imparfaite  (transversale)  en  chaines 
filiformes  (p.  73). 

Zitterthierchen  sind  Monadinen,  welche,  durch  queere  unvollkommene  selbsttheil- 
ung,  bewegte  Gliederfaden  bilden  (p.  74). 

This  family  contained  five  genera :  Bacterium,  Vibrio,  Spirochaeta,  Spirillum, 
and  Spirodiscus. 

BACTERIUM. 

Character :  Animal  e  familia  Vibrioniorum,  divisione  spontanea  in  catenam 
filiformem  rigidulam  abiens. 

Charactere :  Animal  de  la  famille  des  Vibrionides,  prenant  par  la  division  spontan6e 
la  forme  d'un  fil  articu!6  raide  (p.  75). 

Ehrenberg's  Bacterium  was  a  motile  organism,  and  he  also  saw,  or  thought  he 
saw,  an  organ  of  motion,  and  carefully  figured  it. 

Ich  habe  auch  bei  der  starksten  Art  und  Gattung  Bacterium  ein  Beweguugsorgan 
als  einfachen  wirbelnden  Russell  erkant  (p.  74). 

Besonders  erfreulich  war  mir  der  deutliche  Wirbel  am  Vordertheil  der  kleinen 
Korper  im  farbigen  Wasser,  und  eine  angestrengte  Untersuchung  brachte  mir  sogar 
einen  einfachen  fadenartigen  kurzen  Riissel  zu  directer  Anschauung.  Bei  den 
grossten  Formen  [of  B.  triloculare}  hatte  der  Riissel  ^  der  Korperlange,  bei  den 
kleinen  die  Halfte.  Die  Bewegung  der  Thierchen  war  zitternd  und  um  die  Langsaxe 
walzend  (p.  76). 

In  this  species  he  never  saw  more  than  5  nor  less  than  2  or  3  septa.  He  never 
succeeded  in  making  them  take  up  stains,  such  as  indigo,  carmine,  and  india  ink. 
"  Vom  Fortpflanzungsverhaltniss  sind  nur  Ei-?  Kornchen  und  Selbsttheilung,  ein 


NOMENCLATURE   AND    CLASSIFICATIONS.  169 

rein  thierischer  Character,  erkannt"  (p.  74).  At  this  time  his  genus  was  reduced  to 
one  species,  that  with  which  he  started  out,  viz,  B.  triloculare. 

Ganz  sicher  ist  nur  eine  Art  der  Gattung  (p.  75). 

Von  den  im  Jahre  1828  aufgefiihrten  3  Arten  ist  nur  B.  triloculare  als  Stamm  ver- 
blieben,  die  beiden  andern,  B.  stintillans  und  simplex,  sind  unter  diesen  Namen  zur 
Gattung  Monas  gestellt  (p.  77). 

In  the  Vibrionides  stress  is  laid  on  the  incomplete  self-division  by  which  they 
form  motile  jointed  threads  (bewegte  Gliederfaden)  (p.  74).  Bacterium  is  non-flexile 
(unbiegsam) ;  Vibrio  is  flexile  (schlangenfbnnig  biegsam)  (p.  75).  Bacterium  can 
only  swim  straight  ahead.  There  is  a  more  decided  "  Einschniirung  und  grosseren 
Isolirung  der  Einzelthiere  "  in  Vibrio  than  in  Bacterium.  In  both  genera  division 
takes  place  at  a  right  angle  to  the  long  axis. 

Bacterium  triloculare  (fig.  138)  consisted  of  oval  corpuscles  developing  into 
short  cylinders  with  rounded  ends,  2  to  5  times  as  long  as  broad  (usually  3  times), 
having  as  many  transverse  rays  [septa].     It  was  found  in  1820  in  swamp  water  in 
the  oasis  of  Jupiter  Ammon,  in  Libya.     In  1831,  near  Berlin,  Ehrenberg  found  a 
supposedly  similar  organism  which  he  named  Bacterium  articulatum.    Subsequently 
he  says  he  rediscovered  this  organism  in  standing,  "  modri- 
gen "  water  in  a  glass  in  his  room,  and  finding  what   he 
considered  to  be  transition  forms,  he  abandoned  the  latter 
name  and  united  all  of  these  organisms,  whatever  they  may 
have  been,  under  the  name  of  B.  triloculare.     The  state- 
ments  about  the  flagellum,  and  the  figures  of  1838,  appear  to 
have  been  drawn  wholly  from  the  Berlin  forms,  supposed  to  be 

identical  with  the  African  ones.  The  organisms  were  colorless  and  non-flexile  and 
self-division  was  observed.  Contents  very  finely  granular.  Sluggishly  motile,  but 
"  zahlreich  und  deutlich  durcheinander  fahrend."  The  size  of  the  African  form  is 
said  to  be  ^  linie  ;  the  Berlin  forms  were  ^  to  ^  linie.  The  single  "  Thierchen  " 
in  the  Berlin  form  was  one-fifth  as  long,  i.  e.,  ~  linie.  Ehrenberg's  figures  repre- 
sent a  magnification  of  X  290  and  X  1,000  and  are  much  better  than  those  of 
B.  triloculare  in  the  earlier  work.  They  show  a  transversely  2  to  5  septate,  rod- 
shaped  organism,  with  rounded  extremities,  and  bearing  one  polar  flagellum  about 
one-third  the  length  of  the  body.  There  are  no  paired  rods,  or  constrictions  at  any 
of  the  septa,  but  some  of  the  rods  are  slightly  curved.  The  shape  and  septation  of  the 
figures  is  slightly  suggestive  of  some  of  the  drawings  of  de  Bary's  Bacillus  megate- 
rium  (Pilze  Mycet.  u.  Bact,  fig.  194).  They  also  look  somewhat  like  some  of  the 
involution  forms  of  Bacillus  hortulanus  (Phil.  Tr.  Royal  Soc.,  Series  B,  vol.  191, 
pi.  1 6).  Both  of  these  organisms,  however,  have  peritrichiate  flagella.  The  flagel- 
lum resembles  that  on  species  of  Vibrio. 

Cohn's  drawings  of  Bacterium  termo  are  shown  in  fig.  139,  copied  from  his 
"  Beitrage  "  (Bd.  I,  Heft  2,  Tafel  III).  Cohn  did  not  consider  motility  of  any  generic 
value,  and  consequently  paid  no  attention  to  organs  of  motion.  Dallinger  &  Drys- 


*FiG.  138. — Ehrenberg's  Bacterium  triloculare,  showing  flagella.     From  Die  Infusionsthierchen, 
Plate  V.  fig.  I.  i,  2. 


170 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


X.650 

Fig.  139* 


dale's  conception  of  this  organism,  at  a  time  when  the  air  was  full  of  talk  of  Cohn's 
researches,  is  shown  in  fig.  140.     Dallinger  &  Drysdale's  drawings  were  made  from 

unstained  material,  and  there  is  no  doubt  that 
these  expert  microscopists  actually  saw  what  they 
figured,  viz,  a  schizomycetous  organism  provided 
with  one  polar  flagellum  and  belonging  to  the 
family  Bacteriacese.  Dallinger  afterwards  care- 
fully measured  the  diameter  of  the  flagellum  many 
times  over  in  unstained  material,  grown  in  Cohn's 
fluid. 

As  bearing  on  the  question  whether  Ehrenberg 
could  see  the  flagellum  of  an  unstained  bacterium 
with  the  microscopes  at  his  disposal,  it  is  inter- 
esting to  note  Dallinger's  statement  that  Koch  could  not  see  the  unstained  flagellum 
of  Bacterium  termo  because  he  used  "  low-angled  glasses,  which  are  incompetent  to 
that  demonstration."  Another  remark  of  Dallinger  is  also 
pertinent.  "I  have  learned,"  he  says,  "from  experience 
that  there  is  as  great  a  diversity  in  different  individuals  in 
the  sensitiveness  of  the  retina  as  there  is  in  sensitiveness 
of  the  olfactory  or  auditory  nerves." 

The  writer's  own  conception  of  Bacterium  termo  is  shown 
in  fig.  141.  These  organisms  are  green-fluorescent  species 
cultivated  in  Cohn's  solution,  from  water  into  which  beans 
had  been  thrown  in  the  manner  described  by  Cohn.  The 
very  distinct  flagella  were  stained  by  Lowit's  method.  The 
particular  species  from  which  this  was  obtained  did  not 
liquefy  gelatin. 

To  the  writer,  then,  the  genus  Bacterium  is  Bacterium 
(Cohn  emend.),  and  is  based  on  the  morphology  of  the  green-fluorescent  organisms, 

capable  of  growing  in  Cohn's  nutrient  solution  and 
called  by  him  Bacterium  termo.\  It  corresponds 
to  Migula's  genus  Pseudomonas,  for  which  name  it 
should  be  substituted  as  a  proper  generic  name  for 
straignt  or  slightly  curved  Bacteriacese,  motile  by 
means  of  one  to  several  polar  flagella.  It  includes 
most  of  the  yellow  bacteria  and  all  of  the  green-fluor- 
escent bacteria  (vide  Migula's  system,  Bd.  II,  p.  875). 


x? 


X  GOO  ca 
Fig.  I40.t 


(S* 


100 


m  m 


Fig.  1414 


*Fic.  139. — Bacterium  termo:  a,  motile  form;  b,  zoogloese.  After  Cohn.  Untersuchungen  iiber 
Bakterien.  Beitrage  z.  Biol.  d.  Pflanzen,  Bd.  I,  Heft  2,  Plate  III. 

tFic.  140. — Dallinger  and  Drysdale's  conception  of  Bacterium  termo.  See  Dallinger  and  Drys- 
dale  "  On  the  Existence  of  Flagella  in  B.  termo."  Monthly  Micros.  Jour.,  Sept.  i,  1875,  Plate 
CXIII,  p.  105,  figs.  6  and  7. 

JFiG.  141. — The  writer's  conception  of  Cohn's  Bacterium  termo.  Organism  obtained  by  throw- 
ing beans  into  water  and  then  making  a  transfer  from  the  green-fluorescent  liquid  to  Cohn's  solu- 
tion. Stained  by  Lowit's  method.  X  2000. 

gThese  organisms  have  no  necessary  connection  with  Bacterium  termo  Ehrenberg  or  with 
Monas  termo  Muller.  We  shall  never  know  what  these  were. 


NOMENCLATURE   AND    CLASSIFICATIONS. 

We  have  therefore  the  following : 

Bacterium  (Cohn  emend.). 

Type :  The  one-flagellate,  green-fluorescent  schizomycetes,  capable  of  growing 
in  Cohn's  nutrient  solution.  To  these  should  be  added  all  the  morphologically  sim- 
ilar, non -fluorescent  and  yellow  species. 

Synonym  :    Pseudomonas  Migula. 

Among  others  the  following  plant  parasites  belong  here : 

Bacterium  campestre  (Paminel),  B.  pruni  (Erw.  Sm.), 

B.  hyacinthi  Wakker,  B.  vascularum  (Cobb), 

B.  phaseoli  (Erw.  Sm.),  B.  juglandis  (Pierce), 

B.  Steivarti  (Erw.  Sm.),  B.  malvacearum  (Erw.  Sm.). 

These  changes  leave  no  generic  name  for  the  anthrax  organism  and  other  non- 
motile  forms. 

The  writer  wonld  like  to  name  the  anthrax  organism  and  related  forms  in 
honor  of  the  distinguished  man  who  first  pointed  out  the  generic  significance  of 
non-motility  in  this  organism,  but  who  unfortunately  selected  for  it  the  preoccupied 
name  of  Bacteridium.  There  is,  however,  already  a  genus  Davainea  in  helminth- 
ology,  and  it  does  not  seem  wise  to  make  another,  even  in  botany.  Bacteria  are 
now  classed  as  plants,  but  we  do  not  know  what  may  finally  be  done  with  them. 
It  remains,  therefore,  to  adopt  some  old  name,  if  an  unobjectionable  one  can  be 
found,  and  if  not,  to  devise  some  entirely  new  name  for  the  non-motile  bacteria. 
There  are  several  old  names  not  now  in  use,  e.  g.,  Metalader  and  Melanella,  but  so 
far  as  I  have  been  able  to  determine,  none  of  them  were  given  to  organisms  at  all 
resembling  the  anthrax  organism,  and  for  one  reason  or  another  all  imist  be  rejected. 

I  therefore  propose  the  name  Aplanobacter  (from   Greek  words  meaning  without 
motion  and  a  rod),  and  shall  use  it  as  the  generic  name  for  the  anthrax  organism 
called  Bacteridium  by  Davaine,  Bacillus  by  Cohn  and  Fischer,  and  Bacterium  by 
Migula.    Under  Aplanobacter  I  include  all  non-motile  forms  morphologically  similar 
to  the  anthrax  organism  {Bacillus  anthracis  Cohn),  the  latter,  however,  being  taken 
as  the  type  of  the  genus : 

Aplanobacter  nov.  gen.  nom. 

An  unattached,  non-motile,  rod-shaped  organism,  destitute  of  chlorophyll  and 
multiplying  by  fission,  sometimes  forming  threads  of  considerable  length.  The  type 
of  the  genus,  in  the  family  Bacteriacese,  is  that  organism  causing  anthrax  and  most 
commonly  known  in  literature  as  Bacillus  anthracis  Cohn. 

For  the  present  non-sporiferous  forms,  resembling  Aplanobacter  anthracis,  are 
also  included  under  this  genus,  but  if  it  shall  be  decided,  later  on,  that  the  difference 
between  sporiferous  and  non-sporiferous  forms  is  of  generic  significance,  then  the 
latter  may  be  excluded.  This  genus,  as  now  understood,  includes  Aplanobacter 
anthracis  (Cohn)  and  many  other  non-motile  species  called  Bacillus  in  most  books, 
but  Bacterium  by  Migula.  For  a  list  of  the  species  see  Bacterium  (p.  279)  in  Bd. 

II  of  Migula's  "System."     A  few  species  there  given  are  now  known  to  be  motile. 

Forms  related  to  Bacillus  tuberculosis  Koch  and  Bacillus  leprae  Hansen  do  not 
seem  to  belong  with  the  anthrax  organism,  and  some  name  must  be  found  for  these. 


BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 

Lehmann  &  Neumann  have  suggested  Mycobacterium,  and  we  may  use  this  name 
without  in  any  way  committing  ourselves  as  to  the  significance  of  the  branching 
forms.  I  would  include  also  under  it  Bacillus  diphtheria  Loeffler  (Corynebac- 
terium  L.  &  N.).  The  writer  has  not  inquired  critically  as  to  whether  this  is  the 
earliest  available  name  for  this  group,  but  that  of  Sclerothrix,  given  by  Metchnikoff 
in  1888,  is  twice  preoccupied,  and  that  of  Cocothrix,  given  by  Lutz  in  1886,  is  too 
near  the  earlier  Cocothrichium  Link.  In  1889,  in  Saccardo's  Sylloge  Fungorum, 
De-Toni  and  Trevisan  included  these  organisms  under  the  genus  Pacinia  Trevisan, 
but  Trevisan's  original  draft  of  this  genus  included  only  vibrios,  his  type  being 
the  organism  causing  Asiatic  cholera. 

Another  difficulty  is  to  decide  what  name  shall  be  used  for  the  cause  of  Asiatic 
cholera  and  its  relatives.  The  majority,  perhaps,  of  pathologists  and  bacteriologists 
use  the  genus  name  Vibrio.  They  understand  by  it  small  spirally  bent  organisms 
common  in  water  and  possessed  of  one  polar  flagellum  or  rarely  of  several  such 
organs,  the  Vibrio  cholera  being  taken  as  the  type.  Others  call  most  of  these 
organisms  Vibrio,  but  speak  of  Spirillum  cholera.  Others  use  the  two  names 
Vibrio  and  Spirillum  interchangeably.  Others  try  to  escape  the  difficulty  by 
avoiding  Latin  names  altogether,  speaking  in  the  same  article  indifferently  of  "the 
cholera  vibrio,"  "the  cholera  bacterium,"  and  "the  cholera  bacillus."  This  is  the 
case  frequently  in  the  recent  big  monograph  by  Kolle  &  Wassermann.  A  few  per- 
sons, following  Migula,  have  used  Schroeter's  name,  Microspira,  given  in  1886. 
Microspira  is  inadmissible,  according  to  strict  rules  of  priority,  because  Trevisan's 
name  Pacinia  is  one  year  earlier  (1885).  Trevisan's  genus,  although  badly  defined, 
following  Zopf 's  ideas  of  pleomorphism,  is  tied  hard  and  fast  to  the  cholera  organ- 
ism. Apparently  this  name  was  given  without  any  personal  acquaintance  with  the 
organism  named,  but  according  to  current  rules  of  nomenclature  this  makes  no  differ- 
ence. The  choice,  therefore,  is  between  Pacinia  and  Vibrio,  the  one  tied  fast  to  a 
known  species,  but  not  used  by  working  pathologists  or  bacteriologists  since  it 
was  coined,  so  far  as  my  reading  goes,  the  other  in  common  use,  but  a  floating 
name — that  is,  one  which  can  not  be  used  for  bacteria,  and  at  the  same  time  tied 
to  any  definite  species  or  group  of  species  included  in  the  original  draft  of  the  genus. 

Miiller's  genus  Vibrio  was  published  in  1773  in  his  "Vermium  terrestrium 
et  fluviatilium."  It  contained  15  species — bacteria,  eel-worms,  etc.  Other  things 
were  also  afterward  put  into  it  by  Miiller,  e.g.,  diatoms.  We  will  be  content  with 
the  first  draft  of  the  genus.  It  is  described  as  follows :  "  Vibrio.  Most  simple, 
inconspicuous,  terete,  elongate  worms."  The  first  species  is  described  as  follows: 
Vibrio  lineola. 

Vibrio  linear,  hard  to  see.  Danish,  Strseg-strsekkeren.  A  most  minute  animal, 
almost  exceeding  in  smallness  Manas  termo  and  30  times  less  than  Vibrio  bacillus  and 
entirely  different.  A  trembling  motion  of  myriads  of  oblong  and  obscure  points  is 
seen  in  a  single  drop,  or  with  the  highest  magnification  undulatory  movements.  In 
infusions  of  vegetables  it  almost  fills  the  substance  of  the  water  after  many  days. 

The  second  species,  V.  bacillus,  first  obtained  from  hay  infusions,  is  described 
at  a  little  greater  length,  but  not  any  better.  The  third  species  is  a  mixture  of 
nematodes.  The  first  two  species  are  bacteria.  One  other  species  of  schizomycete 
is  described,  viz,  Vibrio  undula.  This  last,  or  what  was  supposed  to  be  it,  together 


NOMENCLATURE   AND    CLASSIFICATIONS.  173 

with  Vibrio  spirillum,  a  subsequent  addition  by  Miiller,  was  removed  by  Ehrenberg 
to  form  his  genus  Spirillum,  which  we  still  retain.  The  eel-worms  were  removed  to 
form  the  genus  Anguillula,  and  the  other  infusoria  were  variously  distributed.  Only 
the  first  two  species  of  the  original  genus  remained  in  Colm's  time,  and  neither  one 
was  used  by  him.  Cohn  used  Vibrio  rugula,  one  of  Miiller's  additions,  for  the  first 
species  under  his  emended  genus  Vibrio,  but  this  has  now  been  put  by  Migula  into 
Spirillum.  The  only  other  member  of  Colm's  genus  Vibrio  (emend.),  V.  serpens,  is 
still  less  like  the  cholera  organism.  Ehrenberg's  figure  of  Vibrio  lineola  Miiller 
(Infusionsth.)  shows  crooked  little  organisms  not  unlike  what  we  now  call  vibrios. 
As  a  general  proposition  the  writer  believes  that  if  a  genus  name  is  to  be 
retained  one  should  be  able  to  tie  it  to  some  definite  type-species,  and  it  ought  to 
be  a  species  put  into  a  genus  when  it  was  first  published,  and  not  one  put  in  after 
the  genus  has  been  emended  out  of  all  recognition.  Of  course,  nothing  can  be 
done  with  Miiller's,  or  Cohn's,  genus  description  of  Vibrio.  If  the  name  is  to  be 
retained  for  any  organisms  whatsoever,  the  description  must  be  made  over  and  the 
name  anchored  to  a  known  species.  Ordinarily  such  a  name  should  be  discarded. 
Under  the  circumstances,  we  may  perhaps  strain  a  point,  make  over  the  genus 
description  in  toto,  and  use  the  name  Vibrio,  as  many  pathologists  have  done,  for 
Koch's  comma  bacillus  and  related  forms.  Logically,  perhaps,  we  should  adopt  the 
strange  Pacinia;  for  convenience  sake  we  may  continue  to  use  the  familiar  Vibrio. 
The  name  Vibrio  is  not  used  by  helminthologists  or  algologists,  and,  if  we  connect 
it  to  the  first  species  described  by  Miiller  under  the  genus,  we  may  anchor  the  name 
to  any  small  motile  species,  without  fear  that  subsequent  researches  will  require 
changes  to  be  made.  This  may  be  done,  because  the  description  of  Miiller's  Vibrio 
lineola,  the  first  species,  is  so  imperfect  that  identification  is  out  of  question ;  the 
name  can  never  be  attached  to  any  morphologically  definite  organism  or  group  of 
organisms  different  from  the  cholera  vibrio,  even  the  gelatinization  of  the  water 
after  many  days  being  probably  enough  due  to  other  bacteria.  The  writer  follows 
Lafar  (ist  ed.),  Alfred  Fischer,  Lehmann  &  Neumann,  et  al.,  and  would  write : 

Vibrio  (Miiller,  Cohn,  emend.).* 

Type  of  the  genus,  Koch's  comma  bacillus. 

Synonyms. — Spirillum  choleree-asiatica  Koch;  Microspira  comma  Schroeter;  Pacinia 
cholera- asiatica  Trevisan . 

Kendall  has  criticized  Migula's  use  of  the  word  Pseudomonas  on  the  ground 
that  he  has  combined  under  it  two  distinct  groups  of  the  family  Bacteriacese,  the 
monotrichiate  and  the  lophotrichiate  forms,  and  because  the  name  implies,  he  says, 
a  relation  to  "  pseudomouads."  The  second  criticism  implies  that  to  be  tenable  a 
name  must  conform  etymologically  to  all  the  facts  in  the  case.  This  is  a  miscon- 
ception. No  one  is  warranted  in  setting  aside  a  generic  or  specific  name  simply 
because  it  seems  inappropriate.  It  is  not  inappropriate,  however,  since  the  first 
species  in  Miiller's  genus  Monas  was  undoubtedly  founded  on  small  bacteria  of 
some  sort.  As  to  the  first  criticism,  that  lies  also  against  my  use  of  Bacterium  and 
requires  a  word.  This  criticism  appears  to  me  not  well  taken,  since  in  the 
Bacteriacese,  as  Migula  first  pointed  out,  there  is  no  such  sharp  distinction 

*According  to  Fischer,  1903,  and  Lehmann  &  Neumann,  1896,  this  emendation  was  made  by  Loeffler. 


174 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


between  the  monotrichiate  and  the  lophotrichiate  forms  as  there  seems  to  be 
among  the  Spirillacese.  Within  the  limits  of  the  same  species,  and  on  the  same 
cover-slip,  forms  may  occur  with  one  flagellum  and  others  with  two  or  more  (see 
fig.  15,  which  is  not  the  only  one  I  might  offer).  The  name  Pseudomoiias  is  of 
earlier  date  than  Fischer's  or  Kendall's  equivalents,  has  priority,  and  can  not  be 
set  aside  on  the  grounds  named.  If  the  reader  is  not  satisfied  with  the  reasons  I 
have  given  for  substituting  the  earlier  name  Bacterium,  then  he  should  continue  to 
use  the  name  Pseudomonas. 

For  the  present,  therefore,  I  follow  Migula's  classification,  except  in  so  far  as 
relates  to  his  use  of  the  words  Microspira,  Pseudomonas,  and  Bacterium. 

The  following  names  should  be  rejected  : 


Acromatium. 

Actinobacter. 

Actinomyces. 

Aerobacter. 

Aethylbacillus. 

Amylobacter. 

Arthrobacter. 

Arthrobactridium . 

Arthrobactrillum . 

Arthrobactrium. 

Ascobacillus. 

Ascobacterium. 

Ascpcoccus.       ,„_ 

Astasia. 

Astrobacter. 

Azotobacter. 

Babesia. 

Bacteridium. 

Bacteriopsis. 

Bactrillum. 

Bactrinium. 

Bollingera. 

Botryomyces. 

Cenomesia. 

Chromatium. 

Clathrocystis. 

Clostridium. 

Clostrillum. 

Clostrinium. 

Coccos. 

Cocobacillus. 

Cocobacteria. 

Cocothrix. 

Cohnia. 

Cornilia. 

Corynebacterium . 

Cryptococcus. 

Cystobacter. 

Dicoccia. 


Diplectridium. 

Diplobacteria. 

Diplococcus. 

Discomyces. 

Dispora. 

Erebonema. 

Ery  throbacillus . 

Erythroconis. 

Fenobacter. 

Gaffkya. 

Gallionella. 

Gliabacteria. 

Gliacoccus. 

Glischrqbacteriuni . 

Gonococcus. 

Granulobacter. 

Gummibacillus. 

Haematococcus. 

Halibacterium. 

Helicomonas. 

Helobacteria. 

Hyalococcus. 

lodococcus. 

Klebsiella. 

Kurthia. 

Lactobacter. 

Lampropedia . 

Leptothrix. 

L,eucocystis. 

Leuconostoc. 

Lineola. 

Macrococcus. 

Megabacterium . 

Megacoccus. 

Melanella. 

Meningococcus. 

Merismopedia. 

Mesobacterium . 

Mesococcus. 


Metallacter. 

Microbacterium . 

Microhaloa. 

Microphyta. 

Microsphsera. 

Microspora. 

Microsporon. 

Microzoa. 

Microzyma. 

Monas. 

Monobacteria. 

Monococcus. 

Mycoderma. 

Myconostoc. 

Mycothece. 

Mycothrix. 

Neisseria. 

Newskia. 

Nitrobacter. 

Nitrosococcus. 

Nitrosomonas. 

Nocardia. 

Nosema. 

Octopsis. 

Ophidomonas. 

Pacinia. 

Paracloster. 

Paraplectrum. 

Pasteurella. 

Pasteuria. 

Pediococcus. 

Perroncitoa. 

Petalococcus. 

Photobacillus. 

Photobacterium . 

Photospirillum. 

Plectridium. 

Pleurococcus. 

Pneumobacillus. 


Pneumococcus. 

Pollendra. 

Proteobacter. 

Proteus. 

Rhabdomonas. 

Rhizobium. 

Saccharobacillus . 

Saccharobacter . 

Schinzia. 

Schuetzia. 

Sclerothrix. 

Sphserococcus. 

Sphserotilus. 

Spirobacillus. 

Spirodiscus. 

Spiromonas. 

Spirulina. 

Sporonema. 

Streblothrichia. 

Streptobacillus. 

Streptobacteria. 

Streptothrix. 

Tetracoccus. 

Thermoactinom  yces . 

Thermobacillus. 

Thermobacteriuni . 

Thioderma. 

Thiosphsera. 

Thiosphserion . 

Torula. 

Tyrothrix. 

Ulvina. 

Urobacillus. 

Urobacter. 

Urocephalum. 

Urococcus. 

Urosarcina. 

Zooglcea. 

Zopfiella. 


PLATE  21. 


Walnut  disease. 

Bacterial  black  spot  of  the  Persian  walnut  (Jualans  rrgia),  more  commonly  known  as  the  English  walnut.  Half-developed  green  fruit*  from  an 
orchard  in  California,  showing  the  badly  spotted  epicarp  ;  spots  due  to  Bacterium  juglandis  (Pieice).  Leaves  and  shoots  are  also  subject  to  this 
disease,  which  has  become  serious  in  Southern  California,  where  large  quantities  of  these  nuts  are  grown  for  market.  The  attacked  parts  are 
conspicuously  blackened  as  if  charred.  The  numerous  small  white  spots  show  the  location  of  group*  of  stomata.  Infection  takes  place  readily 
through  unbroken  tissues. 


NOMENCLATURE   AND    CLASSIFICATIONS.  175 

A  very  few  of  the  preceding  may  perhaps  some  time  make  good  their  claim  to 
be  considered  as  independent  genera.  Many  of  these  names  are  preoccupied  in  this 
group  or  in  other  groups  ;  some  represent  mixtures  ;  others,  purely  physiological 
genera ;  but  some  of  them  may  be  used  within  the  limits  of  genera  to  designate 
special  physiological  groups  whenever  such  use  leads  to  clearness  of  understanding. 

Naegeli,  Beyerinck,  and  Winogradsky  have  studied  especially  the  food  require- 
ments of  bacteria.  Many  others  have,  of  course,  contributed.  Alfred  Fischer  has 
given  a  good  summary  in  the  second  edition  (p.  96)  of  his  "  Vorlesungen."  Follow- 
ing this,  and  considering  them  especially  with  reference  to  their  nitrogen-nutrition, 
the  bacteria  may  be  classified  into  seven  groups  : 

1 .  Paratrophic  bacteria.     The  obligate  parasites,  capable  of  growing  only  on  substrata 

similar  in  composition  to  the  fluids  of  the  host. 

2.  Peptone-bacteria.     Organisms  requiring  peptones  or  albumoses. 

3.  Amido-bacteria.      Organisms  which  also  grow  well  when  their  nitrogen  food  is 

restricted  to  amido-bodies — asparagin,  leucin,  etc. — but  not  able  to  use  ammonia. 

4.  Ammonia-bacteria.     Able  to  take  nitrogen  from  ammonia  compounds. 

5.  Nitrobacteria.     The  denitrifying  organisms.     They  require  organic  carbon  com- 

pounds. 

6.  Nitrous  and  nitrate  bacteria.     The  saltpeter-bacteria.     Nitrates,  nitrites,  or  ammo- 

nia-compounds furnish  the  necessary  nitrogen.     The  carbon  dioxide  of  the  air 
serves  as  their  carbon-food. 

7.  Nitrogen-bacteria.     Organisms  able  to  assimilate  free  nitrogen,  but  only  in  the 

presence  of  organic  carbon  compounds. 

In  1895  Wyatt  Johnston  suggested  that  all  the  important  characteristics  of  a 
species  might  be  recorded  by  numbers  arranged  in  a  definite  order.  Gage  &  Phelps 
and  Kendall  afterward  made  use  of  the  Dewey  numeral  system.  By  this  means 
the  leading  features  of  a  hundred  or  of  five  hundred  organisms  might  be  recorded 
on  a  single  page,  so  as  to  be  very  easily  compared.  Chester  has  modified  this  system 
for  application  within  the  genus  as  follows: 

TOO.  Endospores  produced.  0.002     Acid  without  gas  from  saccharose. 

200.  Endospores  not  produced.  .003     No  acid  from  saccharose. 

10.  Aerobic  and  facultative  anaerobic.  .0001       Nitrates  reduced. 

20.  Anaerobic.  .0002       Nitrates  not  reduced. 

1.  Gelatin  liquefied.  .00001         Fluorescent 

2.  Gelatin  not  liquefied.  .00002        Violet  chromogens. 
o.i  Acid  and  gas  from  dextrose.  .00003        Blue  chromogens. 

.2  Acid  without  gas  from  dextrose.  .00004  Green  chromogens. 

.3  No  acid  from  dextrose.  .00005  Yellow  chromogens. 

.01  Acid  and  gas  from  lactose.  .00006  Orange  chromogens. 

.02  Acid  without  gas  from  lactose.  .00007  Red  chromogens. 

.03  No  acid  from  lactose.  .00008  Brown  chromogens. 

.001  Acid  and  gas  from  saccharose.  .00000  Non-chromogenic. 

According  to  this  scheme  the  formula  for  Bacillus  coli  and  Bacterium  carnpestre 
would  be  respectively  B.  212.11110  and  Bact.  211.33315.  Such  a  system  admits  of 
indefinite  extension,  and  the  reader  can  see  at  a  glance  that,  if  well  worked  out  so 
as  to  include  all  the  more  important  facts,  it  would  be  invaluable  for  unification  of 
methods  and  for  quick,  easy  reference.  Each  group  of  digits  should  include  as 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


many  facts  as  possible.     Kendall,  for  instance,  under  the  gelatin  group  has  also 
included  action  on  dextrose  as  follows  : 


Liquefaction  of 
gelatin. 

Fermentation  of 
dextrose,  gas 
production. 

Acid 
production. 

I. 

Negative. 

Negative. 

Negative. 

2. 

Negative. 

Positive. 

Positive. 

3- 

Negative. 

Negative. 

Positive. 

4- 

Positive. 

Negative. 

Negative. 

5- 

Positive. 

Positive. 

Positive. 

6. 

Positive. 

Negative. 

Positive. 

7- 

Unknown. 

Negative. 

Negative. 

8. 

Unknown. 

Positive. 

Positive. 

9- 

Unknown. 

Negative. 

Positive. 

The  subject  is  now  in  the  hands  of  a  committee  of  the  Society  of  American 
Bacteriologists  for  consideration  and  recommendation,  and  criticisms  are  desired. 
They  may  be  sent  to  Prof.  F.  D.  Chester,  Wilmington,  Del. ;  Prof.  F.  P.  Gorham, 
Providence,  R.  L,  or  to  Erwin  F.  Smith,  Washington,  D.  C. 

VALUE  OF  MORPHOLOGICAL  CHARACTERS. 

Ebb  and  flow,  growth  and  change,  this  is  the  order  of  the  world.  Living  things 
conform  to  a  certain  set  of  conditions  and  we  say  they  are  constant  in  structure 
and  function  because  the  conditions  are  fairly  constant ;  change  the  environment 
too  much  and  they  are  destroyed ;  change  it  essentially,  ever  so  little,  and  the  animal 
or  plant  begins  at  once  to  respond  to  it.  This  is  especially  true  of  simple  uni- 
cellular forms.  We  can  not,  then,  expect  more  than  a  moderate  amount  of  constancy 
in  these  low  forms  of  life.  If  under  slight  changes  of  environment  they  are  fairly 
constant  morphologically,  it  is  all  that  we  can  expect,  and  in  interpreting  all 
descriptions  we  must  make  due  allowance  for  these  slight  changes  which  an  author 
may  not  have  observed. 

There  have  been  two  extreme  views  respecting  the  morphology  of  the  bacteria. 
Be"champ,  Hallier,  Billroth,  and  Zopf  stand  for  one  extreme ;  Koch's  earlier  views 
for  the  other.  To  Hallier  bacteria  were  only  the  developed  plastids  (protoplasmic 
granules)  of  fungi,  and  under  widely  different  forms  we  might  have  the  same 
organism  functioning  at  one  time  as  a  harmless  mold  and  at  another  as  a  micrococcus, 
causing  the  dreaded  cholera  or  some  other  human  or  animal  disease.  Be"champ's 
microzymas  were  granules  or  fundamental  elements  more  minute  than  the  plant  or 
animal  cell,  granules  out  of  which  all  life  developed  and  which  persisted  in  other  forms 
after  the  death  of  the  cells.  To  Billroth  all  ordinary  forms  of  bacteria,  however  dis- 
similar they  might  appear,  were  but  stages  of  one  unique  species,  viz,  his  Coccobacteria 
septica.  Zopf  did  not  carry  his  doctrine  so  far,  but  taught  pleomorphism  as  a  funda- 
mental characteristic  of  the  bacteria.  To-day  an  organism  might  be  a  Micrococcus, 
tomorrow  a  Bacterium  or  a  Bacillus.  Koch,  on  the  other  hand,  insisted  on  the 
fixity  of  forms.  To  him  a  bacillus  was  always  the  same  thing,  and  the  views  of  the 
polymorphists  were  explained  as  the  result  of  errors  in  technique,  the  confounding 
of  entirely  different  things.  Koch's  own  methods  were  exact  and  his  views  had 


PLATE  22. 


L 


Bacterial  black  spot  of  the  walnut. 

A  late  stage  of  the  disease  on  the  nuts.  Photograph  by  Pierce.  Mr.  Pierce,  who  discovered  the  cause  of  this  disease,  has 
demonstrated  50  per  cent  of  the  losses  preventable  by  spraying,  and  is  now  endeavoring  to  obtain  resistant  varieties  by 
hybridizing  and  selection.  The  sum  of  $20,000  was  offered  by  the  walnut  growers  of  California  some  time  ago  for  a 
satisfactory  remedy,  and  recently  the  legislature  of  California  has  appropriated  a  considerable  sum  for  its  investigation. 


NOMENCLATURE   AND   CLASSIFICATIONS.  177 

enormous  weight,  since  he  depended  not  on  mere  assertion,  but  pointed  out  many 
errors  of  fact  and  many  flaws  in  the  reasoning  of  his  antagonists. 

To-day  the  majority  of  bacteriologists  hold  a  sort  of  middle  ground.  Very  few 
are  willing  to  accept  the  views  of  the  old  polymorphists,  but  there  is  a  spirit  of 
rational  inquiry  abroad.  We  know  that  bacteria  are  much  more  responsive  to 
changed  environment  than  was  supposed  by  Koch  and  his  followers  in  the  eighties, 
and  we  are  prepared  to  believe  anything  respecting  their  origin  and  their  poly- 
morphism which  can  gain  the  suffrage  of  the  great  body  of  critical  workers  who 
now  cultivate  this  field,  and  who  at  once  begin  to  investigate  from  all  sides  any  new 
and  strange  statement.  Duplication  of  work,  so  called,  is  not  waste  of  time.*  If 
sharp  criticism  abound,  so  much  the  better.  In  this  way  we  shall  gradually  reach 
a  clearer  understanding  of  these  organisms.  Meanwhile,  let  each  one  cultivate  his 
own  little  field  as  best  he  may,  and,  above  all,  let  him  be  very  sure  of  his  facts  before 
he  publishes. 

There  can  be  no  doubt  that  the  same  organism  sometimes  exists  as  a  long  fila- 
ment in  which  no  septa  are  visible  and  at  other  times  as  a  short  or  nearly  isodia- 
metric  rod,  but  we  are  not  thereby  compelled  to  consider  the  short  form  as  a  Micro- 
coccus,  i.  e.,  as  something  very  different  from  the  long  form.  Physical  conditions 
probably  have  much  to  do  with  bringing  about  these  differences.  Respecting  the 
meaning  of  the  branched  forms,  described  by  so  many  writers,  the  author  is  in  doubt 
and  can  only  wait  for  more  light.  Several  hypotheses  are  open  :  (i)  The  bacteria, 
as  now  understood,  are  not  a  homogeneous  group,  but  consist  of  many  organisms 
of  dissimilar  origin  and  differing  morphologically,  which  will  be  gradually  separated 
out  and  put  into  their  proper  places,  just  as  the  Oosporas  (Streptothrices)  have 
already  been  removed,  leaving  as  jGw-bacteria  a  genuine  residuum  of  morphologically 
similar  forms  ;  (2)  the  bacteria  do  not  any  of  them  represent  a  natural  group,  but 
are  stages  of  various  higher  forms,  just  as  certain  cells,  multiplying  indefinitely  in 
yeast  form,  are  now  known  to  be  conidial  stages  of  the  higher  fungi  (smuts,  mucors)  ; 

(3)  the  branched  forms,  which  come  mostly  in  old  cultures,  or  in  other  crowded 
conditions  where  the  organisms  are  subject  to  the  injurious  action  of  their  own 
by-products  (root-tubercles  of  Leguminosse,  lung-tubercles,  etc.),  are  to  be  regarded 
simply  as  involution  or  degeneration  forms,  and  not  higher  stages  of  development ; 

(4)  the  branchings  are  incomplete  longitudinal  fissions  favored  by  special  chemical 
or  physical  conditions.     Time  will  show  where  the  truth  lies. 

No  harm  will  come  to  any  one  if  all  of  these  perplexing  questions  are  not 
settled  definitely  within  his  own  generation. 

So  far  as  can  be  judged  from  structure  the  bacteria  appeared  in  early  geologic 
ages  (in  coprolites,  decaying  bones,  tree-trunks,  etc.)  in  forms  closely  resembling 
those  now  existing,  but  we  have  very  little  definite  information  as  to  their  origin. 
Probably  they  are  related  to  the  lower  algse  and  of  as  ancient  origin.  On  rela- 
tionship of  the  bacteria  to  Algse,  Fungi,  Flagellata,  and  Myxomycetes,  see  Migula's 
remarks  on  the  systematic  position  of  the  bacteria,  in  his  "System,"  part  I,  page  237. 

*Karl  Pearson  has  recently  stated  that  50  per  cent  of  the  scientific  work  of  the  ipth  century  will 
have  to  be  junked  as  worthless.    In  bacteriology  75  per  cent  would  be  nearer  the  truth. 


178  BACTERIA  IN  RELATION  TO  PLANT  DISEASES. 

VALUE  OF  CULTURAL  CHARACTERS. 

Of  what  worth  are  the  cultural  characters  commonly  mentioned  in  descriptive 
bacteriology  ?  Much  depends  on  the  proper  answer  to  this  question.  There  are 
undoubtedly  two  extreme  views,  neither  of  which  is  correct.  One  investigator 
would  maintain  that  no  dependence  can  be  placed  on  them  ;  another  seems  to  have 
no  suspicion  of  any  source  of  uncertainty.  The  tmth  undoubtedly  lies  somewhere 
between  the  two.  That  great  progress  in  bacteriology  has  come  from  their  use 
must  be  admitted  by  all.  To  cast  doubt  on  everything  already  done  is  only  to 
bring  chaos  back  again.  It  is  wise  to  make  haste  slowly.  No  necessity  exists  for 
making  a  rubbish  heap  of  the  past  before  beginning  one's  own  work.  Old  methods 
should  be  tried  repeatedly,  scrutinized  from  every  standpoint,  and  only  abandoned 
when  they  have  yielded  all  that  can  be  obtained  from  them,  or  when  there  is  some- 
thing distinctly  better  to  take  their  place.  New  methods  should  be  hailed  with 
enthusiasm  only  in  so  far  as  they  have  actually  made  good  their  claim  to  be  genuine 
improvements.  A  great  deal  of  writing  on  bacteriology  is  worthless  because  not 
based  on  well-considered  and  properly  conducted  experiments.  Hypotheses  ad 
libitum,  the  more  the  better ;  but  let  us  not  forget  to  test  each  one  in  the  crucible 
of  experiment,  and  generally  before  publishing,  rather  than  after.  In  other  words, 
give  to  the  world  only  the  well-established  facts.  As  a  means  toward  arriving  at  the 
truth,  let  each  person  not  only  experiment  as  carefully  as  possible,  but  let  him  set 
down  all  the  steps  in  his  procedure,  so  that  others  may  repeat  his  experiments.  Many 
misapprehensions  and  supposed  contradictions  arise  from  the  fact  that  workers  are 
led  to  believe  they  have  exactly  duplicated  another  man's  work  when  they  have  done 
nothing  of  the  kind.  The  temperature  at  which  they  have  worked  has  been  dif- 
ferent, or  some  other  physical  or  chemical  condition,  important  but  not  recognized 
or  not  recorded  by  the  first  writer,  has  been  unlike,  and  the  results  are  not  the 
same.  Bacteria  are  not  so  simple  as  they  appear.  While  monotonous  morphol- 
ogically they  are  complex  in  their  multitudinous  physiological  activities,  and  are 
extremely  apt  to  vary  under  a  slightly  changed  environment.  When  we  repeat  an 
experiment  we  must  know,  therefore,  whether  we  have  preserved  substantially  the 
former  environment.  If  we  have  not,  then  it  should  not  surprise  us  if  the  results 
are  somewhat  different  from  those  we  anticipated. 


c         ~^ 

IT 


A  very  frequent  source  of  error  in  interpreting  descriptions  consists  in  not 
making  sufficient  allowance  for  changes  due  to  slight  variations  in  the  culture-media. 
I  can  perhaps  make  my  meaning  plainer  in  the  following  way :  Let  the  curve  A  B 
represent  all  the  variations  in  color  and  appearance  of  a  given  organism  on  a  given 
medium,  e.  g.,  steamed  potato.  Now,  if  a  worker  describes  his  organism  from  a 


PLATE  23. 


Bacterial  wilt  of  the  cucumber. 

(Introduced  to  illustrate  transmission  of  the  disease  by  insects).  The  central  plant  (variety  Long  green)  wai  inoculated  on  June  17  with  Bacillus 
tracheiphilus  by  the  striped  cucumber-beetle  (Diabrotica  vittata).  As  a  result  the  gnawed  leaves  first  wilted  and  then  the  whole  upper  part  of 
the  plant,  the  vascular  bundles  being  occluded  by  the  sticky  white  slime  of  this  bacillus.  Photographed  July  1 ,  1 905.  About  1-14  natura 
«ze.  The  entire  plant  was  dead  about  two  weeks  later. 


VALUE  OF  CULTURAL  CHARACTERS. 


179 


few  cultures,  he  will  then  in  all  probability  have  covered  only  a  fraction  of  the  curve 
A  B,  let  us  say  between  C  and  D,  and  not  the  whole  curve  of  growth.  If,  now, 
another  worker  should  happen  to  experiment  with  potatoes  capable  of  giving  rise  in 
the  organism  to  phenomena  represented  by  that  part  of  the  curve  lying  between  A 
and  A',  he  would  get  somewhat  different  results  and  yet  this  would  not  prove 
steamed  potato  to  be  a  worthless  culture-medium.  The  only  real  facts  in  the  sup- 
posed case  are  that  neither  person  has  experimented  sufficiently  to  draw  up  a  proper 
description  of  the  characteristics  of  the  given  organism  on  potato.  Let  us  suppose 
we  have  to  do  with  a  yellow  organism,  e.  g.,  Bacterium  phaseoli  and  that  A  to  A' 
represents  a  pale  yellow  growth,  with  no  graying  of  the  potato,  while  D  to  B  repre- 


Fig.  142.* 

sents  a  very  deep  yellow  growth,  with  very  decided  graying  of  the  potato.  The 
cultures  look  like  different  organisms,  but  they  are  not.  The  descriptions  would 
differ.  Neither  account  alone  would  form  a  proper  description  of  the  behavior  of 
this  organism  on  potato,  but  there  should  be  rather  a  combination  of  the  two  and 
of  all  intermediate  stages,  viz — Potato  :  Color  varying  from  pale  to  deep  yellow, 
flesh  of  the  potato  usually  grayed,  but  sometimes  remaining  unchanged,  etc.  The 
same  remarks  apply  to  other  non-synthetic  media. 

*Fic.  142. — Iris-rhizome  rot.  A  dense  sowing  of  the  organism  in  an  agar-plate  culture  after  45 
hours  at  25°  C.  The  buried  colonies  small.  Not  van  Hall's  organism,  which,  as  received  from 
Krai  of  Prague,  is  non-pathogenic  in  my  hands. 


i8o 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


In  case  of  agar  and  gelatin  there  are  numerous  variations  due  to  inadvertent 
changes  in  the  culture-medium,  especially  if  this  is  made  by  students.  The 
media  should  be  made  by  competent,  experienced  persons,  and  then  the  descriptions 
of  the  behavior  of  the  organism  on  it  should  be  broad  enough  to  include  slight 
differences  in  the  aspect  of  the  colonies,  streaks,  and  stabs,  which  often  depend  on 
chemical  and  physical  conditions  within  the  control  of  the  experimenter,  e.  g.,  on 
the  water-content,  on  age  of  the  medium,  amount  of  moisture  in  surface-layers, 
kind  of  peptone,  kind  of  gelatin,  length  of  exposure  and  degree  of  heat  during 
sterilization,  etc.  The  dense  or  thin  sowing  of  the  plate  may  sometimes  make  a 
very  decided  difference  in  the  aspect  of  the  colonies.  Fig.  142  shows  a  densely- 


Fig.  143* 

sown  plate,  the  colonies  round  or  roundish.  Fig.  143  shows  the  same  organism, 
and  from  the  same  set  of  plates,  but  thinly  sown  and  two  days  older.  Here  the 
colonies  are  radiate.  In  case  of  Bacillus  aroidece  when  grown  on  agar-plates, 
near  the  maximum  and  minimum  temperature  limits,  the  surface-colonies  are  round 
even  after  many  days,  but  they  are  promptly  and  strongly  radiate  when  grown  at 
or  near  the  optimum  temperature  (see  figs.  144,  145).  When  very  thin  sowings  of 
this  organism  were  exposed  to  the  high  temperature,  the  colonies  were  also  round. 
It  occurred  to  the  writer  that  the  round  colonies  obtained  on  the  agar-plates 
exposed  in  the  thermostat  at  37°  C.  might  be  due  to  physical  changes  in  the  surface 


*Fic.  143. — Iris-rhizome  rot.     The  same  as  142,  but  sown  thinly  and  kept  for  4  days  at  25°  C. 


VALUE  OF  CULTURAL  CHARACTERS.  l8l 

layers  of  the  agar,  /.  <?.,  to  rapid  loss  of  water,  and  experiments  have  shown  this  to 
be  the  case.  Two  sets  of  Petri-dish  poured  plates  were  made,  inoculating  from  the 
same  culture.  One  set  was  exposed  in  the  open  thermostat  at  37°  C.,  and  these 
developed  round  colonies,  similar  to  those  shown  in  fig.  144.  The  other  set  was 
inclosed  in  the  same  thermostat,  but  inside  of  a  closed  glass  vessel  containing  water. 
The  colonies  on  these  grew  in  radiating  form,  the  same  as  in  a  third  set  of  plates 
exposed  at  30°  C.  This  does  not  account,  however,  for  the  appearance  of  circular 
colonies  at  low  temperatures.  After  twelve  days'  exposure  in  an  ice-box  the  writer 
obtained  the  same  result  as  Townsend ;  the  colonies  were  not  radiate,  but  looked 
like  those  shown  in  fig.  144. 

UNDERGRADUATE  WORK. 

As  a  rule,  the  results  of  this  kind  of  investigation  are  to  be  distrusted.  The 
fresh  ambition  of  students  and  their  delightful  eagerness  to  take  up  hard  problems 
are  sources  of  great  pleasure  to  every  good  teacher.  At  the  same  time  such  students 
must  be  held  back  rather  than  uiged  on,  since  for  the  most  part  they  are  still  unfitted 
to  do  independent  work,  especially  that  which  involves  the  drawing  of  general 
conclusions  from  a  variety  of  experiments.  The  ordinary  training  of  botanical  and 
zoological  laboratories  will  not  fit  the  student  for  specialization  in  pathology  and 
bacteriology.  Skill  in  this  sort  of  work  must  be  ootained  from  consorting  with 
the  professional  pathologist  and  bacteriologist  In  general,  at  the  present  time 
a  well-equipped  modern  laboratory  devoted  to  animal  pathology  is  a  much  better 
place  for  the  plant  bacteriologist  to  learn  methods  than .  even  our  best-equipped 
botanical  laboratories.  One  of  two  alternatives  is  open  to  the  ambitious  student. 
Either  he  must  submit  to  a  long  and  rigorous  course  of  elementary  study  in  a 
bacteriological  laboratory,  under  a  competent  and  critical  teacher,  or  else  he  must 
be  content  to  pick  up  the  general  principles  of  the  science  out  of  books  and 
journals,  with  much  blundering  and  stumbling  in  the  first  years  of  his  study. 
During  this  nursery  period,  if  he  is  jealous  of  his  own  reputation,  he  will  not 
publish  much.  My  experience  has  led  me  to  discount  very  liberally  the  conclusions 
of  student  investigators,  and  I  consider  those  students  very  unfortunate  whose 
teachers  urge  them  into  precocious  publication.  In  many  cases  nothing  could  be 
more  damaging  to  their  own  reputation  as  scientific  inquirers,  or  more  injurious  to  the 
progress  of  science.  Bad  papers  also  react  upon  the  teachers  of  such  students,  who 
can  not  by  any  shift  evade  responsibility.  My  advice  to  teachers  is  to  discourage 
all  students  who  do  not  show  marked  aptitude,  and  to  give  to  those  who  do  show 
signal  ability  the  best  possible  training  in  methods  of  work,  but  to  discourage 
them  from  undertaking  difficult  pieces  of  original  investigation.  The  only  alterna- 
tive is  for  the  teacher  to  follow  their  work  step  by  step  and  assume  joint  responsi- 
bility for  it  in  the  end.  Even  this  latter  course  is  sometimes  risky,  as  the  history 
of  science  shows  very  conclusively. 

After  a  year  or  two  of  careful  work  on  methods,  under  the  watchful  supervision 
of  a  good  teacher,  the  bright  student  will  have  learned  how  to  avoid  many  of  the 
pitfalls  which  beset  his  way,  and,  if  he  has  acquired  a  proper  training  in  other 
directions,  such  as  general  botany,  modern  physics,  chemistry,  the  modern  languages, 


182 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


etc.,  he  may  be  trusted  to  undertake  some  original  research.  Even  when  once  on 
his  feet  as  an  investigator,  my  advice  to  him  would  be :  Try  every  conclusion 
repeatedly  and  make  haste  slowly.  When  he  becomes  uneasy  at  delays,  let  him 
reflect  that  one  really  good  paper  does  much  to  set  an  unknown  worker  on  his  feet 
among  scientific  men,  whereas  one  or  two  hastily  written,  poor  papers  will  injure 
his  reputation  as  an  investigator  more  than  half  a  dozen  good  papers  subsequently 
published  will  suffice  to  repair.  Moreover,  in  this  age  of  enormously  multiplied 
publication  it  is  impossible  to  read  everything,  and  consequently  if  a  writer  wishes 
to  attract  attention  he  must  have  a  commanding  grasp  of  his  subject ;  must  present 


Fig.  144* 

its  leading  features  in  a  clear,  interesting  style ;  must  be  as  brief  as  the  importance 
of  his  subject  will  warrant,  otherwise  his  words  are  certain  to  be  overwhelmed  and 
lost;  and,  finally,  must  publish  in  a  proper  place,  i.  e.,  not  in  some  obscure  "Transac- 
tions" or  in  some  local  journal  with  a  small  circulation.  When  ready  to  publish, 
stop  and  do  your  work  all  over  again  with  more  care.  This  is  my  advice  to  begin- 
ners. In  the  course  of  such  general  revision  the  chances  are  that  many  statements 
will  require  correction  or  modification,  and  some  may  have  to  be  omitted  altogether. 

*FiG.  144. — Colonies  of  Bacillus  aroideae,  circular  when  grown  on  an  agar  plate  at  37°  to  38°  C., 
i.  e.,  at  a  temperature  near  the  maximum.     Photograph  by  Townsend. 


CONSTANCY    OF    CHARACTERS. 


Iii  any  event,  the  student  must  have  a  considerable  body  of  knowledge,  gained 
by  actual  experiment,  before  his  judgment  is  worth  much.  In  the  beginning  he  is 
apt  to  depend  too  much  on  the  constancy  of  organisms  and  is  certain  to  be  misled 
by  names.  To  illustrate :  To  him  all  agar  is  agar  and  all  gelatin  is  gelatin.  Not 
so,  perhaps,  to  the  organism  with  which  he  is  experimenting.  Slight  differences  in 
the  composition  of  a  culture-medium  sometimes  make  considerable  difference  in 
the  growth  and  general  appearance  of  the  bacteria,  and  this  must  be  taken  into 
account.  After  the  student  has  passed  this  stage  of  development  he  can  interpret 
his  results  much  better.  If,  then,  on  some  culture-medium  he  obtains  results  slightly 


Fig.  145* 

different  from  those  already  published  by  some  author,  he  is  not  immediately  driven 
to  suppose  (i)  that  he  has  a  new  species,  or  (2)  that  the  earlier  writer  was  in 
manifest  error.  Other  hypotheses  now  lie  open  to  him.  He  is  dealing  with  a 
living  and  variable  organism,  and  perhaps  the  conditions  in  his  experiment  are  not 
precisely  like  those  to  which  it  was  subjected  by  the  previous  experimenter.  It 
may  also  be  an  organism  which  has  already  varied  into  many  races  having  slightly 
different  peculiarities.  Only  when  full  weight  has  been  given  to  these  possibilities 
is  he  entitled  to  fall  back  on  the  others.  On  the  other  hand,  however,  he  must  not 


FIG.  145. — Colonies  of  Bacillus  aroideae,  radiatc-fimbriate  when  grown  on  an  agar  plate  at  25°  C. 
Photograph  by  Townsend. 


184 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


escape  Scylla  only  to  fall  into  Charybdis.  It  may  be  that  his  organism  varies  in  all 
sorts  of  ways,  but  he  is  by  no  means  to  assume  this.  Every  hypothesis  must  be  tried 
in  the  reducing  fire  of  exact  experiment. 

Probably  the  best  acquirement  a  student  can  get  from  his  years  of  training  is 
a  spirit  of  self-distrust  leading  to  habitual  caution  in  the  drawing  of  conclusions 
and  the  making  of  general  statements.  Such  a  spirit 
will  preserve  him  from  many  foolish  statements  and 
will  enable  him  to  serve  his  generation  to  the  best  of 
his  ability.  He  will  not  go  far,  however,  without  a 
tremendous  earnestness,  an  indomitable  energy,  directed 
in  proper  channels.  Let  him  concentrate  this  energy, 
the  most  priceless  of  all  human  attributes,  and  attack 
specific  problems,  one  after  another  or  a  few  at  a  time ; 
not  all  at  once.  Honesty,  industry,  and  self-reliance, 
tempered  with  the  self-distrust  already  mentioned,  will 
then  carry  him  very  far  on  the  road  he  desires  to  go. 
Finally,  the  student  should  remember  that  the  ideal 
man  of  science,  and  to  a  large  extent  also  the  actual 
man  of  science,  is  a  modest  man,  always  inclined  to  be 
cautious,  always  willing  to  revise  his  conclusions  in  the 
light  of  fresh  evidence,  generally  plain-spoken,  always 
an  enemy  of  shams,  and  never  offended  by  frank  and 
honest  criticism,  preferring  the  white  light  of  truth  to 
the  plaitdits  of  the  multitude. 

A  FINAL  CAUTION. 

Probably  more  mistakes  arise  from  failure  to 
carefully  check  up  the  work  behind  one  than 
from  any  other  source.    What  is  meant  by  this 
can  be  explained  in  a  few  words, 
by  means  of  a  series  of  examples. 

(i)  I  make  subcultures  from 
a  poured-plate  colony.  The  first 
subculture  is  on  slant  agar,  the 
second  is  from  the  agar  into 
beef-broth,  the  third  is  from  the 
beef-broth  into  potato  -  broth, 
and  from  the  latter  I  propose 

to  inoculate  a  plant.     The  in- 

Fig.  146.* 


*FiG.  146. — Apparatus  for  removing  water  from  tissues  with  a  minimum  of  injury.  The  speci- 
men is  placed  on  the  wire  carrier  at  X  in  water.  The  tube  at  the  right  also  contains  water.  Alco- 
hol (95  per  cent)  is  then  poured  into  the  funnel  and  allowed  to  pass  into  the  apparatus  drop  by 
drop.  Its  perfect  diffusion  through  the  water  is  obtained  by  making  the  basal  ends  of  the  carrying 
tubes  flaring  or  funnel-shaped.  By  gaging  the  time  between  drops  the  alcohol  may  be  substituted 
for  the  water,  slowly  or  rapidly,  in  any  desired  time.  About  one-third  actual  size. 


METHODS   OF   WORK.  185 

ference  is  that  this  tube  of  potato-broth,  which  is  only  the  third  remove  from  the 
colony,  contains  a  pure  culture  of  the  organism  with  which  I  started,  but  simple 
observations  of  the  tube,  even  when  coupled  with  a  very  firm  persuasion,  do  not 
assure  me  that  such  is  the  fact.  I  check  the  inference  by  making  plate-cultures  and 
find  in  the  tube  either  (a)  only  the  original  organism ;  (b)  a  mixture  of  two  or  more 
organisms ;  (c)  a  pure  culture  of  some  wholly  different  organism,  which  entered 
during  one  of  the  transfers  as  an  accidental  contamination  and  has  crowded  out  the 
original  organism. 

(2)  A  plant  is  inoculated  from  a  solid  culture  or  fluid  culture  of  a  supposed  para- 
site, and  becomes  diseased.    The  inference  is  that  the  inoculated  organism  has  caused 
the  disease.     I  check  this  inference  by  making  plate-cultures  from  the  interior  of 
the  diseased  tissues  and  find  (a)  great  numbers  of  the  inoculated  organism  in  pure 
culture  and  capable  of  again  producing  the  disease,  which   I  determine  by  actual 
experiment ;  (b)  a  mixture  of  organisms ;  (c)  some  wholly  different  organism ;  (d}  no 
bacteria  whatever. 

(3)  Fermentation-tubes  of  cane-sugar  bouillon  inoculated  with  a  supposedly  pure 
culture  soon  show  clouding  in  the  closed  end,  with  an  abundant  production  of  gas 
and  acid.     The  inference  is  that  these  phenomena  are  due  to  the  presence  of  this 
particular  organism.     I  check  this  inference  by  making  plate-cultures  and  find  (a)  a 
pure  culture  of  the  original  organism;  (b~)  only  an  intruder;  (c)  a  mixture  of  two 
organisms,  in  which  case  both  may  break  up  the  sugar  in  the  manner  described,  or 
only  one  of  them. 

(4)  Drops  of  fluid  containing  a  supposedly  pure  culture  are  dried  on  sterile 
cover-glasses  and  subsequently  put  into  sterile  beef-broth,  which  becomes  clouded. 
The  inference  is  that  the  organism  in  question  has  resisted  the  drying.     I  check 
this  inference  by  making  plate-cultures  from  the  fluid  and  find  (a)  a  pure  culture  of 
the  right  organism  ;  (b)  a  pure  culture  of  some  intruder. 

(5)  The  thermal  death-point  of  an  organism  is  tested  by  inoculating  tubes  of 
beef-bouillon  and  exposing  them  to  a  given  temperature  in  the  manner  already 
described.     Subsequently  the  bouillon  clouds  or  does  not  cloud,  as  the  case  may  be. 
The  inferences  are  that  the  organism  is  killed  or  is  not  killed  by  the  exposure.    The 
first  inference  is  checked  by  having  at  the  same  time  inoculated  other  tubes  of  the 
same  bouillon,  which  have  been  kept  at  room-temperatures,  and  which  (a)  do  not 
cloud,  showing  either  that  the  bouillon  itself  inhibits  growth  or  that  only  dead 
organisms  were  inserted,  i.  e.,  those  from  too  old  a  culture ;  or  (b)  which  cloud  readily, 
showing  that  failure  to  grow  in  the  exposed  tubes  is  actually,  as  it  was  presump- 
tively, attributable  to  the  temperature  of  the  water-bath.     I  check  the  second  infer- 
ence by  making  poured  plates  from  the  clouded  tubes  and  find  (a)  pure  cultures  of 
the  right  organism ;  (b)  pure  cultures  of  some  intruder. 

(6)  A  plant,  which  we  will  designate  as  A,  is  subject  in  the  field  to  a  certain 
disease,  and  this  disease  is  readily  reproduced  under  experimental  conditions,  using 
pure  cultures  of  a  given  microorganism.     A  related  plant,  which  we  designate  as  B, 
is  subject  in  the  field  to  a  similar  disease.     A  microscopic  examination  shows  sim- 
ilar lesions  associated  with  a  morphologically  similar  organism,   and   Petri-dish 


l86  BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 

poured  plates  indicate  the  presence  of  a  physiologically  similar  organism  in  both 
plants.  The  first  inference  is  that  the  two  diseases  are  caused  by  the  same  organism. 
A  test-experiment  is  now  instituted,  viz,  one  or  two  varieties  of  B  are  inoculated 
with  the  organism  obtained  from  A,  but  these  do  not  contract  the  disease.  An 
easy  second  inference  now  is  that  we  are  dealing  with  two  distinct  diseases.  This 
may  be  perfectly  correct,  but  it  is  not  established  by  the  experiment.  Owing  to  an 
oversight,  plants  of  A  were  not  inoculated  at  the  same  time  and  in  the  same  man- 
ner as  B,  to  serve  as  checks,  and  consequently  we  are  not  assured  as  to  the  virulent 
nature  of  our  culture — it  may  have  been  dead,  or  non-virulent,  or  the  wrong  organ- 
ism. Check-plants  should  have  been  inoculated.  Assuming,  however,  that  this  was 
done,  and  that  A  promptly  contracted  the  disease  while  B  remained  unaffected,  it  is 
not  yet  certain  that  the  disease  in  the  two  plants  is  due  to  different  organisms.  The 
question  of  individual  and  varietal  resistance  to  disease  may  have  entered  to  com- 
plicate results.  To  eliminate  this  possible  source  of  error  a  greater  number  of 
varieties  of  B  should  be  tested  with  a  larger  number  of  individuals  in  each  variety. 
Cross-inoculations  should  also  be  made,  i.  e.,  numerous  varieties  and  individuals  of  A 
should  be  inoculated  with  the  organism  isolated  from  B. 

Enough  has  been  said  to  show  the  ordinary  method  of  work.  All  inferences 
should  be  carefully  confirmed  by  frequent  poured-plate  cultures  in  Petri  dishes,  by 
cultivations  on  the  media  which  have  been  found  to  give  most  characteristic  results, 
and,  finally,  by  frequent  inoculations  into  the  host-plants.  In  case  of  unexpected  or 
striking  results  it  is  always  safe  to  determine  whether  they  can  not  be  obtained  in 
the  absence  of  the  assumed  cause. 

These  methods  involve  an  almost  endless  amount  of  drudgery,  but  they  are 
fundamental  to  any  large  success  in  the  domain  of  pathology,  and  those  who  are 
desirous  of  winning  a  shining  reputation  without  much  labor  are  advised  to  culti- 
vate some  easier  science.  For  those  who  are  really  in  earnest,  who  do  not  mind 
hard  work,  and  who  have  acquired  the  requisite  training,  no  field  affords  greater 
opportunity  for  brilliant  and  useful  work  than  that  of  plant  pathology. 


FORMULA. 


When  not  stated  the  solids  are  reckoned  in  grains  and  the  fluids  in  cubic  cen- 
timeters.    Water  is  understood  when  no  particular  solvent  is  mentioned. 

STAINS. 
GENERAL  AND  MISCELLANEOUS. 


Alcoholic  Solutions  of  Anilin  Stains. 
These    should    be    saturated    solutions,    made 
preferably  with  Griibler's  stains  and  absolute  al- 
cohol.   In  well-stoppered  bottles  they  keep  in- 
definitely. 

Watery  Solutions  of  Anilin  Dyes. 

These  do  not  keep  long  and  must  be  made  up 
fresh  each  time.  If  made  directly  from  the  dry 
powder  or  crystals,  rather  than  from  the  alcoholic 
solution,  the  resulting  fluid  should  be  passed 
through  filter  paper  before  using.  Watery  solu- 
tions are  usually  made  by  adding  the  alcoholic 
solution  to  distilled  water  in  any  strength  de- 
sired. Usually  a  few  drops  of  the  alcoholic 
solution  to  5  or  10  cc.  of  water  is  sufficient. 

Anilin  Water. 

Anilin  water  is  made  by  shaking  thoroughly 
one  part  of  anilin  in  20  parts  of  distilled  water 
and  filtering  it  clear  by  passing  one  or  more 
times  through  filter  paper  moistened  with  water. 
It  should  be  prepared  fresh  each  time.  Anilin, 
known  also  as  anilin  oil,  is  a  colorless,  oily- 
looking  fluid.  It  oxidizes  to  a  brown  color  if 
exposed  to  the  air,  and  it  should  therefore  be 
kept  in  a  close-stoppered  bottle  in  the  dark. 
The  brown  fluid  is  still  usable,  at  least  for  some 
purposes. 

Ziehl's  CarboI-Fuchsin. 

Fuchsin  (basic) I 

Absolute  alcohol 10 

Carbolic   acid    (5   per   cent   sol.    in 

water)    100 

The  fuchsin  should  first  be  dissolved  in  the 
alcohol  and  then  the  two  fluids  mixed.  A  pow- 
erful and  much-used  stain. 

Ehrlich's  Anilin-Water  Gentian   Violet. 

Alcoholic  solution  of  gentian  violet 

(saturated)    5 

Anilin  water 100 

This  should  be  used  as  soon  as  prepared.  It 
does  not  keep  well. 


Flexuer's  Anilin  Gentian  Violet. 

Anilin  oil 2 

Alcohol,  95  per  cent 5 

Saturated  alcoholic   (absolute)   solu- 
tion of  gentian  violet 8 

Distilled  water So 

Mix  well  and  filter. 

Ehrlich-Weigert  Anilin  Methyl  Violet. 
Alcoholic  solution  of  methyl  violet 

(saturated)   n 

Absolute  alcohol 10 

Anilin  water 100 

Does  not  keep  well. 

Anilin  Fuchsin. 

Prepared  in  the  same  way  as  Ehrlich's  anilin 
gentian  violet 

Ziehl-Nielson's  Stain. 

Used  chiefly  as  a  means  for  identifying  tuber- 
culosis. The  cover-glass  bearing  the  specimen 
is  floated  for  3  to  7  minutes  on  carbol-fuchsin 
which  is  heated  until  steam  begins  to  appear. 
It  is  then  washed  in  distilled  water,  plunged  into 
10  per  cent  nitric  or  sulphuric  acid  long  enough 
to  decolorize  (a  very  short  time).  It  is  then 
passed  through  60  per  cent  alcohol  for  a  few 
seconds  (just  long  enough  to  remove  the  stain 
from  the  background),  washed  thoroughly  in 
water,  dried,  and  mounted  in  balsam.  The 
cover-glass  preparation  may  be  obtained  also  by 
dropping  some  of  the  stain  upon  it  and  holding 
it  over  the  flame.  This  method  is  more  eco- 
nomical of  stain  and  time  and  less  mussy  than 
the  preceding. 

Friedlaender's  Stain. 

This  has  been  used  so  far  mostly  for  identify- 
ing the  tubercle  organism  in  sputum.  It  is 
made  as  follows :  A  few  drops  of  carbol-fuchsin 
are  placed  on  the  prepared  cover  (which  has 
been  gently  flamed)  and  heated  over  a  flame 
until  the  fluid  steams.  The  cover  is  then 
washed  in  distilled  water,  and  plunged  for  a  half 

187 


1 88 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


minute  or  so  into  acid  alcohol  (c.  p.  nitric  acid 
5  cc.,  80  per  cent  alcohol  100  cc.).  It  is  then 
washed  in  water,  stained  about  5  minutes  (for 
contrast)  in  an  aqueous  solution  of  methylene 
blue,  dried,  and  mounted  in  cedar  oil  or  balsam. 

LoefRer's  Alkaline  Methylene  Blue. 
Alcoholic     solution     of     methylene 

blue  (saturated) 30 

Caustic  potash i  )  JQO 

Distilled  water 10,000  j 

This  fluid  retains  its  valuable  properties  for  a 
considerable  time  and  is  an  excellent  stain. 

Kuhne's  Carbol-Methylene  Blue. 

(1)  Methylene  blue i-S 

Absolute  alcohol 10.0 

(2)  After  triturating  the  above  in  an  agate  or 
porcelain  mortar,  or  in  a  watch  glass,  add  grad- 
ually 100  cc.  of  water  containing  5  per  cent  car- 
bolic acid.     Methylene  blue  is  not  the  same  as 
methyl  blue.     (See  Pregl,  Bibliog.,  XIV.) 

Gram's  Stain. 

This  is  a  method  of  differential  bleaching 
after  a  stain.  The  cover-glass  preparations  or 
sections  are  passed  from  absolute  alcohol  into 
Ehrlich's  anilin  gentian  violet  or  into  a  watery 
solution  of  methyl  violet,  where  they  remain 
i  to  3  minutes,  except  tubercle  bacilli  prepara- 
tions, which  remain  commonly  12  to  24  hours 
(Gram).  They  are  then  placed  for  i  to  3  min- 
utes (occasionally  5  minutes)  in  iodine  potas- 
sium iodide  water  (iodine  crystals  I,  potassic 
iodide  2,  water  300),  with  or  without  first  wash- 
ing lightly  in  alcohol.  In  this  they  remain  i  to  3 
minutes.  They  are  then  placed  in  absolute  alco- 
hol until  sufficiently  bleached,  after  which  they 
are  cleared  in  clove  oil  and  mounted  in  Canada 
balsam.  By  this  method  the  stain  is  removed 
from  some  kinds  of  bacteria  and  not  from 
others. 

Too  much  confidence  must  not  be  placed  in 
this  method,  since  in  some  cases  the  removal  or 
non-removal  of  the  stain  from  the  organism  de- 
pends on  the  length  of  exposure  to  the  iodine 
water.  It  would  be  better,  therefore,  to  expose 
all  for  the  same  period,  e.  g.,  2  minutes. 

Gabbett's  Stain. 

Used  mostly  for  tubercle  bacteria  in  sputum. 
Stain  first  with  carbol-fuchsin,  then  place  the 
cover-glass  for  i  to  2  minutes  in  acid  methylene 


blue  (methylene  blue  2  grams,  25  per  cent  sul- 
phuric acid  water  100  cc.).  When  washed  in 
water  and  dried  it  may  be  mounted  in  cedar  oil 
or  in  balsam.  The  ordinary  bacteria  of  sputum 
are  decolorized ;  the  tubercle  organism  retains 
the  red  stain. 

The  Ehrlich-W eigert  Stain. 

Used  for  detecting  the  tubercle  organism  in 
sputum.  The  prepared  cover  is  floated  face 
down  on  anilin  methyl  violet,  which  is  heated 
until  steam  rises.  After  2  to  5  minutes  on  this 
hot  stain  plunge  for  a  few  seconds  into  acidu- 
lated water  (i  part  nitric  acid,  3  parts  distilled 
water),  then  wash  for  a  few  seconds  in  60  per 
cent  alcohol,  and  afterward  thoroughly  in 
water.  For  a  contrast  stain  the  cover  may  be 
placed  for  5  minutes  in  a  saturated  aqueous  so- 
lution of  vesuvin.  It  is  then  washed  in  water, 
dried,  and  mounted  in  balsam. 

Bacteria  which  hold  the  stain  after  such  treat- 
ment are  sometimes  called  "  acid-fast "  bacteria. 

Flcmming's  Triple  Stain. 
The  slide  is  first  placed  in  (i). 

(1)  Safranin  O   (saturated  alcoholic 

solution)   50 

Distilled  water 50 

Anilin  water 5 

After  washing  in  water,  it  then  goes  into  (2). 

(2)  'Saturated    aqueous    solution    of 

gentian  violet 50 

It  is  then  washed   in  water  and  passed  into  (3). 

(3)  Aqueous     solution     of    orange     G, 

strong  or  weak    (generally  about 
one-half  saturated). 

The  slide  is  then  washed  quickly  in  95  per 
cent  alcohol,  dehydrated,  cleared,  and  mounted. 

Pregl's  Method. 
(See  '91  Pregl,  Bibliog.,  XIV.) 
Nicolle's  Methods. 
(See  '95  Nicolle,  Bibliog.,  XIV.) 

Benda's  Iron  Haematoxylin. 
Mordant  the  sections  for  several  hours  in   i 
part   of   the   following   ferric   solution*   diluted 
with  2  parts  of  water: 

Ferrous  sulphate 80 

Water 4° 

Sulphuric  acid 15 

Nitric  acid 18 


•This  solution,  known  to  the  German  Pharmacopoeia  as  Liquor  ferri  sulphuric!  oxydati  and  to  the  U.  S.  P.  as  Liq. 
f.  tersulphatis  or  sol.  persulphate  of  iron,  keeps  indefinitely.  It  is  made  as  follows  :  Heat  in  a  flask  on  a  water-bath 
until  fluid  is  brown  and  clear,  and  a  drop  diluted  with  water  is  no  longer  colored  blue  by  potassium  ferricyauide, 
evaporate  in  a  tared  porcelain  capsule  to  100  parts,  add  a  little  water  and  evaporate  again.  Repeat  the  dilution  and 
evaporation  until  the  hot  fluid  is  free  from  the  odor  of  nitric  acid.  Finally  dilute  to  a  weight  of  160  parts. 


STAINS. 


189 


Wash  the  sections  in  distilled  water  and  then 
in  tap-water.  Stain  (until  very  black)  in  water 
containing  I  per  cent  haematoxylin.  Differen- 
tiate in  30  per  cent  acetic-acid  water  with  care- 
ful watching,  or  in  more  dilute  acid,  or  in  very 
dilute  (i  :  20)  liquor  ferri,  if  it  is  to  be  followed 
by  acid  fuchsin  as  a  contrast  stain.  (Verhandl. 
d.  Anat.  Gessellsch.,  1893,  Jena,  Gustav  Fischer.) 

Heidenhain's  Iron  Haematoxylin. 
Mordant  the  sections  from  one-half  hour  to 
12  hours  in  a  2.5  per  cent  watery  solution  of  iron 
alum  (ammonio-ferric  sulphate  (NH4)i  Fe2 
(SO<)i  dissolved  cold).  This  salt  comes  in 
violet  crystals.  Yellow  or  green  crystals  should 
be  rejected.  Rinse  well  in  water.  Stain  in 
water  containing  0.5  per  cent  of  haematoxylin. 
Rinse,  expose  again  to  the  iron-alum  solution, 
watching  the  differentiation  under  the  micro- 
scope, the  examination  being  made  in  tap-water. 
When  properly  differentiated  wash  15  to  45  min- 
utes in  running  water.  Dehydrate,  clear  only 
with  xylol,  and  mount  in  xylol  balsam.  The 
mordant  does  not  keep  indefinitely,  but  is  said 
to  retain  its  properties  for  some  time  (Dodge). 


Schaffner's  Safraiiin  Picro-nigrosin. 

The  slides  are  stained  for  a  few  minutes  in 
anilin-safranin  made  as  follows : 

1 i )  Anilin  water 50 

Saturated     alcoholic     (95     per 

cent)  solution  of  safranin 50 

Rinse  quickly  in  water.  They  are  then  stained 
in  (2). 

(2)  Distilled  water 100 

Picric  acid  thoroughly  dissolved 

in  the  above i 

Then  add  nigrosin i 

Rinse  in  water,  wash  rapidly  in  95  per  cent  al- 
cohol, dehydrate,  and  mount  in  balsam. 

Malachite  Green. 

For  plant  tissues,  as  well  as  animal  tissues, 
this  may  be  used  as  a  contrast  stain,  follow- 
ing carbol-fuchsin.  It  is  dissolved  in  anilin 
(i  :  1000)  and  used  fresh;  generally  the  ex- 
posure to  it  should  be  for  only  a  very  brief 
period,  »'.  e.,  i  to  3  minutes.  If  not  fresh,  much 
longer  exposures  are  required. 


CLEANING  COVER-GLASSES  FOR  FLAGELLA  STAINS. 

Van  Ermengem  recommends  boiling  in  a  mixture  of  water  100  cc.,  concen- 
trated sulphuric  acid  60  cc.,  potassium  bichromate  60  grams.  The  covers  are 
afterward  thoroughly  washed,  first  in  water  and  then  in  absolute  alcohol.  They 
are  set  on  edge  and  dried  under  a  bell-jar. 

Loeffler  recommends  heating  covers  in  concentrated  sulphuric  acid.  They  are 
then  washed  in  distilled  water  and  put  into  alcohol-ammonia,  from  which  they  are 
wiped  with  a  very  clean  linen  cloth. 

Covers  cleaned  in  the  ordinary  way  may  be  freed  from  fat  by  passing  them 
through  a  Bunsen  flame  immediately  before  using. 

FLAGELLA   STAINING. 

There  is  no  easy  road  to  success.  Some  of  the  common  stones  of  stumbling 
are  (i)  oily  or  otherwise  dirty  covers  ;  (2)  cultures  unsuited  either  by  age  or  by 
composition  of  the  medium  ;  (3)  the  casting  off  of  flagella  on  dilution  or  during 
the  slow  drying  of  the  fluid  on  the  cover  ;  (4)  an  uneven  or  too  copious  distribution 
of  the  organisms  on  the  cover ;  (5)  imperfect  mordanting ;  (6)  excessive  mor- 
danting ;  (7)  understating  ;  (8)  overstating  ;  (9)  precipitates  on  the  cover-glass 
during  some  stage  in  the  process. 

If  clean  covers  are  used,  if  the  bacteria  are  derived  from  young  moist  agar  cult- 
ures, if  a  very  small  quantity  of  such  culture  is  put  into  a  large  drop  of  well  aerated 
water,  or,  better,  into  a  test-tube  or  watch-glass  containing  5  or  10  cc.  of  water,  and  a 
tiny  quantity  of  this  dilution  is  taken  on  a  platinum  needle  and  deftly  swept  over 
the  whole  cover ;  or  if  the  needle  is  touched  to  the  bacterial  fluid  and  then  touched 


190 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


to  one  or  more  parts  of  a  large  drop  of  water  on  the  cover,  which  is  then  put  into 
the  thermostat,  so  that  it  shall  dry  quickly  and  yet  allow  time  for  various  rods  to 
swim  free  from  the  tangle  of  their  fellows  ;  if  the  mordanting  is  thorough,  but  not 
excessive,  and,  finally,  if  there  is  no  unforeseen  mishap  in  the  subsequent  staining, 
good  preparations  of  many  organisms  are  easily  secured.  Others  have  given  the 
writer  much  trouble  {Bacillus  amylovorus  is  one  of  the  worst),  and  the  only  con- 
clusion he  has  been  able  to  reach  is  that  the  bacteria  vary  greatly  in  their  response 
to  flagella  stains.  Sometimes  well-cleaned  covers  give  trouble  and  then  the  surface 
of  the  glass  itself  is  at  fault  (V.  A.  Moore),  and  covers  of  another  origin  should 
be  tried. 

Young  agar  cultures  are  usually  preferred,  i.  e.,  those  not  over  twenty-four  hours 
old,  but  good  results  may  occasionally  be  obtained  from  much  older  cultures  and 
also  from  other  media.  In  case  flagella  are  to  be  stained  from  fluid  cultures  by 
ordinary  methods,  the  medium  in  which  the  organism  is  grown  must  be  very  dilute, 
and  there  must  usually  be  an  additional  extensive  dilution  on  the  cover,  or  before 
putting  thereon,  to  avoid  a  dense  ground  stain.  Bouillon  contains  too  many  fine 
particles,  but  Zettnow  has  found  a  way  to  stain  from  bouillon  (Bibliog.,  XII). 

Recently  the  writer  has  obtained  very  good  results  with  several  organisms  by 
growing  them  for  some  days  in  10  cc.  of  distilled  water  to  which  2  or  3  drops  of 
Uschinsky's  solution  was  added.  Covers  were  prepared  directly  from  this  solution. 
Others  have  reported  good  success  by  transfer  of  agar-grown  organisms  to  consider- 
able water  in  a  watch-glass  or  test-tube  and  incubating  this  in  the  thermostat  for 
some  hours  before  preparation  of  the  covers.  Others  have  recommended  the  use  of 
filtered,  sterile,  hydrant  water  instead  of  distilled  water.  Nearly  every  worker  has 
some  favorite  stain.  The  writer  prefers  Van  Ermengem's. 


Loefier's  Flagella  Stain. 

(ij 'Mordant: 

Solution  of  tannin  (20  per  cent  in  water) ....  10 
Saturated  (cold)  aqueous  solution  of  ferrous 

sulphate*  5 

Saturated  alcoholic  solution  of  basic  fuchsin.     I 

(2)  Stain:  Carbol-fuchsin. 

(3)  Corrective  solutions:   (a)  I  per  cent  solu- 
tion of  caustic  soda;   (&)  a  sulphuric-acid  solu- 
tion of  equivalent  strength. 

V.  A.  Moore's  Flagella  Stain. 

This  is  a  slight  modification  of  Loeffler's. 

(1)  Mordant: 

Tannic  acid  (20  per  cent  in  water) 10 

Ferrous  sulphate  (cold  saturated  water  solu- 
tion)        5 

Basic  fuchsin  (saturated  alcoholic  solution)  .     i 

(2)  Stain :  Ziehl's  carbol  fuchsin. 

Use  the  mordant  fresh,  and  filter  each  time 
before  using.    Fix  the  film  on  the  cover-glass 


by  passing  it  quickly  face  up  twice  through  the 
open  flame,  or  by  exposing  for  5  or  10  minutes 
to  a  temperature  of  120°  to  140°  C.,  the  better 
way.  A  few  cubic  centimeters  of  the  mordant 
are  now  placed  in  a  wide  test-tube  (i  inch),  the 
cover  is  lowered  into  it,  and  the  mordant  is 
heated  over  a  flame  until  steam  rises.  It  is 
then  removed  from  the  flame  and  the  cover  is 
allowed  to  remain  in  the  hot  fluid  5  or  10 
minutes,  with  occasional  gentle  shaking,  after 
which  it  is  drawn  to  the  mouth  of  the  tilted 
tube  by  means  of  a  hooked  platinum  wire  set 
into  a  glass  rod.  After  thoroughly  rinsing  under 
the  tap  or  in  a  fine  stream  of  water  the  cover 
is  lowered  into  the  stain  (held  in  another  wide 
test-tube),  where  it  is  heated  until  steam  rises 
(i  to  3  minutes).  One  should  know  from  be- 
ginning to  end  which  side  of  cover  bears  the 
bacterial  film.  The  cover  is  drawn  to  mouth  of 
tube  by  means  of  a  hooked  platinum  wire.  Some- 
times i  per  cent  sol.  NaOH  (l/2  cc.)  may  be 
added  to  mordant  with  advantage. 


•The  iron  oxide  may  be  removed  from  the  solution  of  ferrous  sulphate  by  passing  it  through  a  filter  paper. 
Selected  crystals  should  be  used. 


FLAGELLA    STAINS. 


191 


Fischer's  Flagella  Stain. 

This  is  a  slight  modification  of  Dr.  Corner's, 
which  is  itself  a  modification  of  Loeffler's. 

(1)  Mordant: 

Dry  tannin 2 

Water  20 

Solution  ferrous  sulphate  (1:2)....     4 
Concentrated  alcoholic  solution  basic 

fuchsin  i 

Filter.  This  mordant  will  keep  for  several 
weeks.  Drop  it  on  the  cover  and  heat  over  a 
gentle  flame  until  steam  rises ;  then  continue  one- 
half  minute  longer  without  boiling.  Wash. 
Treatment  with  alcohol  is  unnecessary. 

(2)  Stain :  Place  on  the  cover  a  few  drops  of 
concentrated   watery  solution  of  basic   fuchsin. 
Heat  slowly,  so  that  steam  rises  after  about  i 
minute.    Then     expose     for     one-half     minute 
longer,  so  that  the  stain  boils  up  once  or  twice. 
Wash,  dry,  and  mount.     Fischer  says  tannin  ab- 
sorbs moisture   readily,  and  advises  keeping  it 
in  a  desiccator. 

Bunge's  Flagella  Stain. 

(1)  Mordant: 

Tannin  20,  water  100 30 

Liquor  ferri  sesquichlorati i 

Water    20 

Saturated     watery     solution     basic 

fuchsin    5 

This  must  ripen  some  weeks  by  exposure  to 
the  air  in  a  flask  loosely  plugged  with  cotton. 

(2)  Stain :  Carbol-fuchsin. 

Expose  to  the  filtered  mordant  5  minutes, 
using  heat  if  necessary.  Wash  and  stain. 

Van  Ermengem's  Flagella  Stain. 
This  is  made  as  follows : 

(1)  Mordant: 

Osmic  acid  (2  per  cent  water  solu- 
tion)       50 

Tannin  (10  to  25  per  cent  in  water) ..  100 
Four  drops  of  glacial  acetic  acid  may  be  added 
to  this. 

(2)  Silver  bath:  0.25  to  0.5  per  cent  nitrate  of 
silver  dissolved  in  distilled  water  in  a  very  clean 
bottle. 

(3)  Reducing  and  strengthening  bath : 

Gallic  acid 5 

Tannin  3 

Fused  sodium  acetate  (some  books 

say  fused  potassium  acetate) 10 

Distilled  water 3SO 

The  flamed  cover-glass  (it  may  be  unflamed) 
is  first  covered  with  the  mordant  for  one-half 


hour,  or  if  in  a  thermostat  at  50°  C.  for  5  or  10 
minutes.  The  mordant  is  then  carefully  re- 
moved by  thorough  washing  in  water,  alcohol 
(some  say  absolute  alcohol),  and  water.  The 
cover  (film  side  up)  is  now  put  into  the  silver 
bath  (a  few  cubic  centimeters  in  a  small,  per- 
fectly clean  beaker  or  watch-glass)  for  a  few 
seconds,  during  which  it  is  gently  agitated. 
Without  rinsing,  it  is  put  next  into  a  few  cubic 
centimeters  of  the  reducing  solution  and  gently 
agitated  until  the  fluid  begins  to  blacken.  It  is 
then  washed  in  water  and  examined.  If  not 
stained  deeply  enough  the  cover  is  returned  to 
the  silver  bath,  then  once  more  passed  through 
the  reducing  bath.  It  is  finally  dried  and 
mounted  in  balsam.  All  the  dishes  must  be 
scrupulously  clean.  The  fluids  must  not  be  con- 
taminated by  the  fingers  nor  by  dipping  iron  or 
steel  instruments  into  them. 

Kuntze  has  suggested  some  improvements. 
(See  Centralb.  f.  Bakt,  I  Abt.,  Bd.  XXXII, 
1902,  pp.  555-560.) 

PMeld's  Flagella  Stain. 

1 i )  Mordant : 

Tannic  acid,  10  per  cent  aq.  sol 10 

Corrosive  sublimate,  sat.  aq.  sol 5 

Alum,  sat.  aq.  sol 5 

Carbol-fuchsin 5 

Heat  on  cover  until  steam  rises;  keep  at  this 
temperature  i  minute;  then  wash,  dry,  and 
stain. 

(2)  Stain : 

Alum,  sat.  (cold)  aq.  sol 10 

Gentian  violet,  sat.  ale.  sol 2 

Kendall,  in  Journ.  Applied  Micr.,  Vol.  V, 
1902,  p.  1836,  says  this  has  proved  a  very  satis- 
factory stain  to  himself  and  his  associates. 

Another  formula  is  given  as  follows : 

(1)  Saturated     (cold)    aqueous    solution    of 

alum  10 

Saturated   alcoholic   solution   of   gentian 
violet   i 

(2)  Tannic  acid   (tannin) i 

Distilled  water 10 

A  mixture  of  these  two  fluids  is  put  on  the 
flamed  cover,  which  is  held  over  the  flame  and 
gently  heated  until  nearly  ready  to  boil.  The 
cover  is  then  put  aside  for  i  minute,  after  which 
it  is  washed  in  water,  dried,  and  mounted  in  bal- 
sam. If  the  mixed  mordant-stain  is  filtered  be- 
fore using,  it  is  best  to  stain  a  second  time  for  a 
moment  in  anilin-water  gentian  violet. 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Lowit's  Flagella  Stain. 

Lowit's  modification  of  Loeffler's  flagella  stain 
consists  in  substituting  a  copper-tannin  mordant 
for  one  of  iron-tannin.  It  is  made  as  follows : 

(l)  Mordant: 

Distilled  water 10 

Tannin   2.5 

Dissolve  and  filter  through  two  thicknesses  of 
filter  paper,  then  add : 

Saturated  solution  copper  sulphate...  5 
Saturated  ale.  sol.  basic  fuchsin I 

Filter  as  before. 

Covers  are  exposed  to  this  mordant  20  seconds 
to  3  minutes  without  heating.  Wash  thor- 
oughly. 

(2)  Stain :  Expose  cover  to  Ehrlich's  anilin 
water  gentian  violet  I  to  5  minutes.  Wash  thor- 
oughly in  water,  and  if  this  is  not  sufficient 
plunge  for  a  moment  into  50  per  cent  alcohol  or 
into  acid  alcohol  (i  drop  of  0.3  per  cent  HC1 
alcohol  in  3  to  4  cc.  of  60  per  cent  alcohol).  The 
mordant  and  stain  must  be  made  up  each  time. 
If  the  mordant  has  been  in  use  for  some  hours 
so  that  an  oxidation  film  has  formed  on  its  sur- 
face, it  is  well  to  stop  and  remove  this  by  filtra- 
tion. 

Sclavo's  Flagella  Stain. 

Sclavo  exposes  some  minutes  in  the  mordant 
(tannin  I,  water  50,  alcohol  50)  ;  washes  in  aq. 
dest. ;  exposes  some  minutes  in  50  per  cent 
phospho-tungstic  acid ;  washes  carefully  in  aq. 
dest. ;  stains  3  to  5  minutes  in  gently  warmed 
anilin-water  fuchsin;  washes,  dries,  and  mounts 
in  Canada  balsam.  Some  kinds  are  not  stained 
by  this  method. 

Bowhill's  Flagella  Stain. 

This  author  used  two  solutions  made  as  fol- 
lows: 

(1)  Orcein  I 

Absolute  alcohol 50 

Distilled  water 40 

(2)  Tannic  acid   (tannin) 8 

Distilled  water  (hot) 40 

Equal  parts  of  (i)  and  (2)  are  mixed  and 
filtered.  Bacteria  from  a  fresh  agar  culture  are 
suspended  in  boiled  distilled  water.  This  sus- 
pension is  allowed  to  stand  5  minutes.  Drops 
are  taken  and  spread  and  dried  on  clean  covers, 
which  are  then  taken  in  the  fingers  and  fixed 
over  the  flame.  They  are  now  floated,  film  down, 
on  the  mordant  (gently  warmed)  for  from  10 
to  15  minutes,  then  washed  in  water  and  dried. 


Ehrlich's  anilin-water  gentian  violet  is  now 
dropped  on  the  cover,  and  this  is  heated  over 
the  flame  until  steam  appears.  The  preparation 
is  then  washed,  dried,  and  mounted  in  xylol 
balsam. 

Subsequently  Bowhill   modified   the  above  as 
follows,  using  the  orcein  itself  as  a  stain: 
(i) 'Saturated    solution    of   orcein   (al- 
lowed to  ripen  about  10  days). 
(2)  20  per  cent  solution  of  tannin  dis- 
solved in  hot  water. 

The  stain  is  prepared  by  taking  of  No.  i,  15 
cc. ;  No.  2,  10  cc. ;  and  distilled  water,  30  cc. 
After  mixing,  the  fluid  should  be  filtered.  Sub- 
sequent bleaching  of  the  preparation  should  be 
avoided.  (Hyg.  Rundsch.,  VIII  Jahrg.,  1898, 
pp.  ii  and  105.) 

Hinterbcrgcr's  Method  for  Flagella. 
OSee  'oo  Hinterberger,  Bibliog.,  XII.) 
Night  Blue  Stain  for  Flagella. 
('See  '99  Morton,  Bibliog.,  XII.) 

Zettnow's  Method  for  Flagella. 

Zettnow  fixes  with  formalin,  mordants  with 
tartrate  of  antimony  and  tannin,  and  stains  with 
gold  or  silver. 

The  fixing  is  done  by  taking  bacteria  from  a 
fresh  agar  or  bouillon  culture  and  adding  them 
to  water.  They  are  then  killed  by  the  addition 
of  4  per  cent  formalin.  The  fixed  bacteria  set- 
tle after  a  day  or  two,  when  the  sediment  con- 
taining them  is  pipetted  out  and  washed,  first 
in  i  per  cent  formalin  water  and  finally  in  pure 
water.  The  cloudy  water  is  spread  and  dried 
on  clean  covers,  and  when  these  have  been  fixed 
by  gentle  heat  they  are  ready  for  the  mordant. 
The  mordant  is  made  as  folows : 

(1)  Tannin   5 

Distilled  water too 

Flask  and  heat  to  35°  or  40°  C.  in  the  water- 
bath. 

(2)  A  solution  of  tartrate  of  antimony  (i  gram 
dissolved    in  water  in  a  test-tube)  is  added  drop 
by  drop  to  solution  (i),  with  shaking  until  the 
precipitate  which  forms  is  not  redissolved.     It 
is  then  filtered.    The  filtered  mordant  should  be 
strongly  opalescent,  but  not  cloudy  or  opaque  to 
transmitted  light.     This  is  said  to  be  a  perma- 
nent universal  mordant,  and  one  which  does  not 
cause  precipitates  on  the  cover.     It  is  used  hot 
(70°  to  80°  C.)  for  5  or  10  minutes.     The  cover 
is  then  washed  and  gilded  or  silvered.     After- 
ward the  image  may  be  intensified  if  desired. 


FLAGELLA    STAINS. 


193 


The  gold  method  consists  of  placing  on  the 
mordanted  cover  4  or  5  drops  of  an  aqueous 
solution  of  neutral  gold  chloride  (i  :2ooo). 
This  is  then  heated  until  steam  is  given  off 
freely.  If  the  mordanting  has  been  sufficient 
there  will  be  a  deposit  of  metallic  gold  on  the 
bacteria. 

Hugh  Williams'  Method. 

(Copied  from  Mallory  &  Wright's  Patholog- 
ical Technique.) 

This  is  a  modification  of  van  Ermengem's 
method  along  the  lines  of  the  modification  of 
Hinterberger  and  others.  It  has  been  adopted 
by  Dr.  Hugh  Williams  after  a  large  experience 
with  various  methods  in  the  laboratory  of  the 
Massachusetts  General  Hospital. 

The  method  is  capable  of  giving  black  bac- 
teria and  flagella,  with  little  or  no  precipitate. 
The  method  is  as  follows : 

(1)  Cover  the  cover-glass  with  a  mordant  con- 
sisting of — • 

Alumnol,*  i  per  cent  solution...   I  part 
Osmic  acid,  2  per  cent  solution. .   i  part 

Tannin,  20  per  cent  solution 3  parts 

Shake  the  mixture,  add  three  drops  of  glacial 
acetic  acid,  and  again  shake. 

(2)  Apply  the  mordant  less  than  one  minute 
without  heating.     Wash  thoroughly  in  water. 

(3)  Cover    the    preparation,    during    about    i 
minute,  with  a  i  per  cent  solution  of  silver  ni- 
trate  to   which   sufficient   ammonium    hydroxid 
has  been  added  to  keep  the  silver  in  solution.-j- 

(4)  Wash  in  water. 

(5)  Wash  with  0.6  per  cent  solution  of  sodium 
chlorid. 

(6)  Flood  the  preparation  with  a  30  per  cent 
solution    of    ammonium    hydroxid,    and    imme- 
diately wash  in  water. 

('/)  Apply  a  few  drops  of  Ortol  photographic 
developer.  (The  directions  for  making  up  this 
developer  come  with  the  Ortol.) 

(8)  Wash  in  water. 

(9)  Cover  with  a  i  per  cent  solution  of  gold 
chlorid  during  a  few  seconds. 

(10)  Wash  in  water  and  apply  Ortol  developer 
for  a  few  seconds. 

(u)  Wash  in  water  and  cover  with  a  i  per 
cent  solution  of  mercuric  chlorid  for  a  few  sec- 
onds. 

(12)  Wash  in  water. 

(13)  Apply  Ortol  developer  for  a  few  seconds. 


(14)  Wash  in  water  and  repeat  the  application 
of  chlorid  of  gold,  the  washing,  and  the  applica- 
tion of  the  developer  two  or  more  times.  Be- 
tween the  various  applications  of  the  chlorid  of 
gold  the  preparation  should  be  inspected  with  a 
high,  dry  lens  to  determine  the  progress  of  the 
staining.  This  is  readily  done  by  placing  the 
cover-glass,  charged  side  upward,  on  a  slide.  In 
this  way  the  process  of  impregnation  with  gold 
may  be  controlled;  for  the  flagella,  if  stained, 
may  be  easily  seen  with  the  high-power  dry  lens. 

The  preparation  is  very  conveniently  held  dur- 
ing the  process  in  cover-glass  forceps.  The 
washing  is  best  done  in  a  small  stream  of  water 
from  a  faucet.  The  various  solutions  are  con- 
veniently applied  from  dropping-bottles. 

It  will  be  seen  that  the  process  consists  essen- 
tially in  the  impregnation  of  the  flagella  with 
silver,  followed  by  intensification,  in  the  photo- 
graphic sense,  with  mercury  and  gold.  The  ob- 
ject of  the  application  of  the  sodium  chlorid  and 
ammonia  is  to  remove  the  excess  of  silver  com- 
pounds which  adhere  to  the  surface  of  the  cover- 
glass  in  spite  of  washing.  This  excess  of  silver 
compounds  is  chiefly  responsible  for  the  precipi- 
tates which  appear  on  the  preparation  after  the 
intensification.  In  spite  of  the  application  of 
the  sodium  chlorid  and  ammonia  solutions, 
some  precipitate  will  occur  if  the  intensification 
is  pushed  too  far.  On  this  acount  it  is  advisable 
to  observe  the  progress  of  the  intensification 
under  the  microscope,  as  above  indicated. 

Although  this  method  may  appear  compli- 
cated, in  practice  it  requires  but  a  few  minutes 
to  stain  a  preparation. 

Duckwall's  Method. 

Streaks  are  made  on  2  per  cent  agar  in  Petri 
dishes  from  young  growths  in  bouillon.  Sus- 
pensions are  made  in  water  according  to  nature 
of  organism,  the  motile  bacteria  being  divided 
into  six  groups  for  staining  purposes.  Pigments 
and  slime  are  removed  by  shaking  with  chloro- 
form. Cover-glasses  must  be  absolutely  clean. 
Mordant  must  be  used  only  when  fresh;  dye 
must  be  fresh  and  used  warm  (Loeffler's  stain)  ; 
streaks  on  cover-glass  should  not  be  confluent. 
Fix  without  injury  to  flagella;  stain  without 
overheating;  wash  in  alcohol  and  water  without 
breaking  film;  clear  in  xylol;  mount  in  xylol 
balsam  without  previous  examination.  For  de- 
tails respecting  method  of  making  suspensions 
see  The  Canner  and  Dried  Fruit  Packer,  Vol. 
XX,  Feb.  16,  1905,  p.  23. 


*Farhwerke  vorm.  Meister  I.ucius  u,  Briining,  Hochst  a.  M.,  Germany. 

fWorkers   in  the  Bureau  of  Animal   Industry  U.  S.  Department  of  Agriculture   state  that  too  great  an  excess  of 
ammonia  In  the  silver  nitrate  may  interfere  with  the  working  of  the  method. 


194 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


CAPSULE  STAINS. 


Ribbert's  Method  of  Staining  Capsules. 

Water 100 

Alcohol  50 

Glacial  acetic  acid 12.5 

Warm  and  add  dahlia  to  saturation.  The 
covers  are  barely  touched  to  this  stain,  and  are 
then  washed  in  water.  The  cover  may  then  be 
mounted  in  glycerin  or  balsam.  The  stain 
keeps  well.  If  the  cover  is  left  on  the  stain  too 
long  the  capsule  becomes  deep  blue  and  can  not 
be  distinguished  from  the  body  of  the  organism. 

Friedlaender's  Capsule  Stain. 
(See  '85  Friedlaender,  Bibliog.,  XIII.) 
Richard  Muir"s  Capsule  Stain. 

1 i )  Mordant : 

Mercuric  chloride  (sat.  aq.  sol.) 2 

Tannin  (20  per  cent  in  water) 2 

Potash  alum  (sat.  aq.  sol.) 5 

The  dried  films  are  mordanted  2  minutes. 

They  are  then  washed  in  water,  in  alcohol,  and 

again  in  water. 

(2)  Stain :  Carbol-fuchsin  2  to  3  minutes,  with 
gentle  heat. 

Wash  with  water;  re-mordant  2  to  3  minutes; 
wash  again  with  water. 

(3)  Counter-stain:    Methylene   blue    (sat.    aq. 
sol.)  2  minutes.     Bleach  in  methyl  alcohol,  clear 
in  xylol. 


Welch's  Capsule  Stain. 

Fix  in  glacial  acetic  acid.  After  a  few  sec- 
onds pour  off  the  acid  and  flood  with  anilin- 
water  gentian  violet;  repeat  this  operation  until 
all  acid  is  removed ;  wash  and  examine  in  salt 
solution  (0.85  to  2  per  cent). 

Kaufmann's  Method. 

Stain  2  hours  at  35°  C.  in  Loeffler's  methylene 
blue;  wash  in  water  containing  caustic  potash 
or  soda  (i  :  1500),  dry;  expose  2  minutes  in  Vi 
per  cent  silver  nitrate;  wash  again  in  the  alka- 
line water ;  counterstain  30  seconds  in  basic 
fuchsin  water  (i  cc.  sat.  ale.  sol.  in  20  cc.  aq. 
dest.)  ;  expose  again  to  the  alkaline  water  (sec- 
onds) ;  dry  and  mount.  Best  adapted  to  demon- 
stration of  capsules  in  fresh  tissues.  The  bac- 
terial body  is  blue  and  the  capsule  red. 

Moore's  Night-blue  Capsule  Stain. 

(See  '99  Moore,  Bibliog.,  XIII.) 
Boni's  Method. 

Mix  white  of  I  egg,  glycerin  50  cc.,  formalin 
2  drops;  shake  well  and  filter.  The  bacteria 
are  placed  in  a  drop  of  this  fluid,  spread,  and 
heated  until  the  glycerin  has  entirely  evaporated. 
.Stain  20  to  30  seconds  in  carbol-fuchsin,  wash 
in  water,  dry  and  counterstain  4  to  6  minutes 
in  Loeffler's  methylene  blue,  wash  in  water,  dry, 
and  mount  in  Canada  balsam.  The  background 
is  red,  body  of  organism  blue,  and  periphery 
colorless.  (See  'oo  Boni,  Bibliog.,  XIII.) 


SPORE  STAINS. 


Mauser's  Spore  Stain. 

Pass  cover-glass  quickly  three  times  through 
flame.  Drop  on  carbol-fuchsin  and  heat  for  5 
minutes  over  flame,  renewing  the  stain  as  it  boils 
away.  Nearly  decolorize  in  dilute  sulphuric  or 
acetic  acid  (5  per  cent).  Wash  very  thoroughly. 
Counterstain  with  a  dilute  watery  solution  of 
methylene  blue  or  with  Loeffler's  methylene  blue 
(Festschrift  fur  Zenker). 

Mailer's  Spore  Stain. 
(See  '91  Moeller,  Bibliog.,  XI.) 
Neisser's  Spore  Stain. 

The  cover  is  floated  on  hot  anilin-fuchsin  for 
an  hour.  The  temperature  should  be  near  the 
boiling  point.  The  cover-glass  is  then  washed 
in  water  and  decolorized  in  acid  alcohol  (i  part 
hydrochloric  acid,  3  parts  alcohol).  Care  must 
be  taken  not  to  expose  to  the  acid  alcohol  too 


long;  otherwise  the  color  will  be  removed  from 
the  spores  also.  The  cover  is  now  stained  for 
contrast  in  a  saturated  aqueous  solution  of 
methylene  blue. 

Fiocca's  Spore  Stain. 

The  prepared  cover  is  placed  in  a  watch-glass 
or  test-tube  containing  20  cc.  of  10  per  cent  am- 
monia (water  solution)  and  10  to  20  drops  of 
alkalin  methylene  blue  or  other  alkalin  solu- 
tion of  anilin  color.  Then  the  fluid  is  heated 
to  the  giving  off  of  steam  and  left  for  3  to  15 
minutes.  It  is  now  passed  for  a  moment  ( ?) 
through  20  per  cent  nitric  or  sulphuric  acid,  then 
thoroughly  washed  in  water  and  stained  for  con- 
trast, if  desired,  in  an  aqueous  solution  of  vesu- 
vin,  malachite  green,  or  safranin  (saturated?). 
(See  '93,  Fiocca,  Bibliog.,  XL) 

(For  other  methods,  e.  g.,  Foth's,  Klein's, 
Aujeszky's,  see  Bibliog.,  XI.) 


FORMULA. 


195 


NON-SYNTHETIC  CULTURE  MEDIA. 


Standard  Peptonized  Beef-Bouillon. 

Standard  peptonized  beef-bouillon  is  made  as 
follows :  To  500  grams  of  finely  minced  lean 
beef  add  1,000  cc.  of  distilled  water.  The  solu- 
ble parts  may  be  removed  from  the  meat  by  al- 
lowing the  water  to  stand  on  it  for  24  hours  in 
the  ice-chest  or  for  I  hour  in  the  water  bath  at 
55°  C.  The  writer  prefers  the  second  method. 
Then  boil  for  60  minutes  either  in  the  steamer 
or  in  a  covered  dish.  Filter  through  a  clean 
cloth,  using  pressure  (meat-press),  cool,  and 
remove  fat  by  filtering  through  S.  &  S.  filter 
paper;  make  up  to  1,000  cc.  by  addition  of  more 
water;  then  add  I  per  cent  Witte's  peptonum 
siccum  and  0.5  per  cent  c.  p.  sodium  chloride. 
Steam  one-half  hour,  filter,  cool,  titrate,  add  re- 
quired alkali,  steam  again  for  one-half  hour, 
filter,  pipette  into  tubes  or  flasks,  and  autoclave 
or  heat  for  a  minimum  time  in  the  steamer. 
Plugs  should  be  well  made  and  fit  tightly; 
glassware  should  be  scrupulously  clean.  For 
some  purposes  both  the  peptone  and  the  salt 
may  be  omitted.  A  greenish  bouillon  indicates 
insufficient  boiling,  and  will  usually  throw  down 
some  additional  vexatious  precipitate  when 
heated  in  the  test-tubes.  Other  meats  may  be 
substituted  for  beef,  and  other  peptones  for 
Witte's.  Meat-extracts  are  not  recommended. 
Such  extracts  usually  contain  resistant  spores. 
Media  which  have  been  steamed,  or  boiled  in  an 
open  dish,  are  better  for  many  bacteria  than 
those  which  have  been  sterilized  in  the  autoclave. 

(For  additional  obse'rvations  on  proper  steril- 
ization see  Culture  Media,  p.  29.) 

Dunham's  Solution. 

Distilled  water 1,000 

Witte's  peptonum  siccum 10 

C.  P.  sodium  chloride 5 

First  recommended  by  Dr.  Ed.  K.  Dunham,  of 
New  York. 

Standard  Nutrient  Agar. 

To  1,000  cc.  of  standard  beef-bouillon  add  10 
grams  of  agar-flour,  steam  one-half  hour,  cool 
to  58°  C.;  add  whites  of  two  eggs  (beaten  thor- 
oughly and  neutralized  to  litmus  by  dilute  hy- 
drochloric acid)  and  thoroughly  mix  with  the 
bouillon;  steam  I  hour,  filter  hot  through  S.  &  S. 
paper  which  has  been  thoroughly  warmed  with 
boiling  distilled  water.  Use  two  or  three  fun- 
nels. That  which  remains  unfiltered  after  a 
reasonable  time  must  be  reheated  and  put 


through  a  fresh  filter  paper.  Sometimes  all  can 
be  got  through  a  second  filter  paper  without  re- 
heating. Some  advise  filtering  in  the  autoclave 
or  in  the  steamer,  but  the  writer  has  not  found 
that  necessary,  and  in  recent  years  has  also 
abandoned  the  hot-water  funnel. 

Clear  agar  may  be  obtained  also  by  filtering 
through  absorbent  cotton,  and  some  prefer  this 
to  filter  paper. 

In  preparing  agar  from  the  "  slender  kanten  " 
or  the  "  square  kanten,"  snip  fine,  soak  in  the 
bouillon  15  minutes,  and  then  heat  on  the  sand 
bath  i  hour  at  110°  C.  or  in  the  autoclave  45 
minutes  at  105°  C.  From  this  point  proceed  as 
before. 

Long  heatings  in  the  autoclave  at  110°  C.,  or 
shorter  heatings  at  higher  temperatures,  are  apt 
to  brown  the  agar,  and  should  be  avoided  care- 
fully, as  this  renders  the  medium  less  service- 
able for  the  growth  of  bacteria.  Agar  which  has 
been  properly  superheated  filters  readily.  One 
per  cent  agar  made  from  the  agar-flour  does  not 
require  to  be  heated  on  the  sand-bath  or  in  the 
autoclave,  but  filters  satisfactorily  after  steam- 
ing for  an  hour  at  100°  C. 

After  the  agar  has  been  tubed  it  may  be  ster- 
ilized, if  it  does  not  contain  sugars,  by  one 
steaming  in  the  autoclave  for  10  minutes  at  110° 
C.,  or  by  short  steamings  in  the  steam  sterilizer 
at  100°  C.  on  three  successive  days. 

To  those  who  are  dependent  on  the  agar-strips 
and  do  not  have  access  to  an  autoclave,  Schutz's 
method  may  be  recommended  as  very  good. 
The  writer  formerly  made  large  use  of  this.  It 
consists  in  heating  the  agar  very  hot  in  a  mini- 
mum quantity  of  water  or  bouillon  before  add- 
ing the  bulk  of  the  fluid.  (See  p.  34  and  Bib- 
liog.,  XVI.) 

Agar  which  has  been  soaked  in  5  per  cent 
acetic  acid  for  an  hour  or  two  before  adding  it 
to  the  bouillon  also  enters  into  solution  thor- 
oughly and  filters  well  after  a  short  boiling.  The 
acid  must  first  be  removed  completely  by  wash- 
ing in  running  water  for  some  hours  under  a 
mosquito-net  or  a  piece  of  gauze. 

Unfiltered  agar  does  well  enough  for  certain 
fungi,  and  for  lazy  people,  but  the  agar  used  for 
delicate  work  in  bacteriology  should  be  as  clear 
as  the  bouillon  from  which  it  is  made.,  i.  e.,  per- 
fectly free  from  cloudiness  and  precipitates. 
Sufficient  caustic  soda  is  usually  added  to  the 
agar  to  render  it  +  15  of  Fuller's  scale.  Other 
degrees  are  useful,  e.  g.,  +  10,  o,  —  10,  etc. 


196 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Litmus  Lactose  Agar. 

To  1,000  cc.  of  ordinary  agar,  preferably  that 
made  up  with  bouillon  free  from  muscle-sugar, 
add  10  grams  of  c.  p.  lactose  and  20  cc.  of  a  sat- 
urated (water)  solution  of  c.  p.  (lime-free)  blue 
litmus. 

Hesse  and  Niedntr's  Nutrient  Agar  for  Water 
Bacteria. 

Distilled  water 980 

Nahrstoff  Heyden   (an  albumose)       7.5 

Agar-agar  12.5 

This  agar  is  said  to  be  the  most  suitable  me- 
dium for  the  bacteriological  examination  of 
water.  It  gives  a  much  larger  number  of  colo- 
nies than  ordinary  agar.  It  requires  no  neu- 
tralizing. The  poured  plates  are  counted,  ac- 
cording to  Dr.  Robin,  on  the  gth  or  roth  day. 
Chromogenic  species  are  brilliantly  colored. 
(Zeitschr.  f.  Hygiene,  Bd.  XXIX,  pp.  454-462. 
See  also  Am.  Jour.  Pharm.,  Vol.  LXXVI,  p. 
112.) 

Glycerin  Agar. 

To  each  1,000  cc.  of  ordinary  agar  add  50  cc. 
of  Schering's  c.  p.  twice-distilled  glycerin. 

Standard  Nutrient  Gelatin. 
To  1,000  cc.  of  sterile  standard  peptonized 
beef-bouillon  add  100  grams  of  best  quality  gela- 
tin. Soak  2  hours  at  room-temperature.  Then 
steam  5  minutes,  cool,  titrate,  add  the  necessary 
alkali,  steam  30  minutes,  filter  through  S.  &  S. 
paper  washed  with  sterile  boiling  hot  water,  tube 
at  once,  and  heat  in  the  steamer  on  three  suc- 
cessive days  15  minutes,  10  minutes,  and  5  min- 
utes, respectively,  at  100°  C.  Do  not  autoclave, 
and  carefully  avoid  long  heatings  in  the  steamer. 
Have  all  the  glassware  sterile  and  the  fluids 
sterile  and  sufficiently  boiled  to  begin  with. 
Very  acid  gelatin  should  be  avoided.  The  very 
best  English,  French,  and  German  gelatins 
should  be  used.  +  10  or  +  15  is  a  good  degree 
of  alkalinity  for  many  purposes.  Zero  of  Ful- 
ler's scale  is  also  useful.  See  remarks  on  gela- 
tin (p.  30). 

Blood  Serum. 
(See  p.  48.) 

Plant  Juices. 
(See  p.  41.) 

Solid  Vegetable  Substances. 
(See  page  39.) 

Milk. 
(See  p.  46.) 


Litmus  Milk. 

Litmus  milk  is  prepared  from  fresh  milk 
which  has  been  passed  through  a  separator  (cen- 
trifuge), or  from  milk  which  has  stood  18  or  20 
hours  at  20°  C.  and  has  had  the  cream  removed 
by  skimming  and  filtration.  To  each  100  cc.  of 
this  milk  is  added  2  cc.  of  a  saturated  solution  of 
high-grade,  lime-free,  blue  litmus  (litmus  I 
gram,  water  15  cc.).  This  gives  a  lavender  color 
of  just  the  right  degree,  which  reddens  distinctly 
under  the  action  of  acids,  and  blues  with  the 
development  of  alkalies.  The  milk  selected 
should  not  titrate  more  than  +  16  with  phe- 
nolphthalein  and  caustic  soda.  A  good  quality 
often  gives  +  10  to  +  14.  High  readings  de- 
note the  excessive  multiplication  of  lactic  acid 
bacteria.  Such  milks  frequently  coagulate  on 
steaming,  and  are  not  suitable  for  culture-media. 
After  adding  the  litmus  water  the  milk  should 
be  pipetted  in  10  cc.  portions  into  cotton-plugged 
test-tubes  and  heated  in  streaming  steam  (100° 
C.)  for  15  minutes  on  each  of  4  successive  days. 

This  is  a  very  useful  culture  medium.  Every 
organism  should  be  tested  in  it.  All  milk  used 
for  culture  media  should  be  centrifuged,  if  pos- 
sible, immediately  after  milking,  and  secured  at 
once  for  the  laboratory.  Three  steamings  are 
then  sufficient.  Milk  offered  for  sale  in  cities  is 
frequently  more  than  48  hours  old  and  often  con- 
tains from  3,000,0000  to  6,000,000  bacteria  per 
cubic  centimeter.  Such  milk  is  not  fit  for  labo- 
ratory use. 

Nutrient  Starch  Jelly. 

The  writer  makes  this  as  follows:  To  10  cc. 
portions  of  modified  Uschinsky"s  solution,  or  of 
the  ordinary  solution  (glycerin  omitted  or  not,  as 
desired),  is  added  i  gram  of  clean  aseptic  potato 
starch.  This  is  rubbed  up  in  the  slanted  fluid. 
The  test-tubes  are  then  very  tightly  plugged  to 
avoid  loss  of  water  and  placed  carefully  in  a 
blood-serum  oven  or  in  the  top  of  an  Arnold 
steam  sterilizer  with  the  vents  open,  where  they 
are  heated  for  2  hours  on  each  of  5  successive 
days  at  85°  C.  to  93°  C.  If  water  is  lost  during 
the  heating  it  must  be  made  up,  using  a  sterile 
pipette.  Potato  starch  is  prepared  in  the  labora- 
tory (p.  50)  with  care  in  the  washing  and  dry- 
ing, so  as  to  avoid  retention  of  other  substances 
than  starch  and  the  multiplication  of  resistant 
(spore-bearing)  bacteria,  which  interfere  with 
the  sterilization.  (See  Proc.  Am.  Asso.  Adv. 
Sci,  1898,  Vol.  XLVII,  p.  4".) 


FORMULA. 


I97 


SYNTHETIC  CULTURE  MEDIA. 


Pasteur's  Culture  Fluid. 

Ammonium  tartrate 10 

Ashes  of  yeast 10 

Rock  candy 100 

Distilled  water 1,000 

Dissolve  cold. 

Naegeli's  Nutrient  Solution. 

Calcium  chloride o.i 

Magnesium  sulphate .2 

Dipotassium  phosphate i.o 

Ammonium  tartrate 10.0 

Distilled   water 1,000.0 

Cohn's  Nutrient  Solution. 

Distilled   water 1,000.0 

Acid  potassium  phosphate 5.0 

Magnesium  sulphate 5.0 

Neutral  ammonium  tartrate 10.0 

Potassium  chloride 0.5 

(De  Bary,  p.  86,  Vorles.  ii.  Bact.,  2  Auflage.) 
Raulin's  Culture-Fluid. 

Distilled  water 1,500.00 

Granulated  cane  sugar 70.00 

Tartaric  acid 4.00 

Ammonium  nitrate 4.00 

Ammonium  phosphate .60 

Potassium  carbonate .60 

Magnesium  carbonate .40 

Ammonium  sulphate .25 

Zinc  sulphate .07 

Ferrous  sulphate .07 

Potassium  silicate .07 

Prasmowski's  Culture-Fluid. 

Dipotassium  phosphate 5.0 

Magnesium  sulphate 5.0 

Ammonium  carbonate 5.0 

Calcium  chloride .5 

Distilled  water 1,000.0 

Dissolve  cold.  Any  desired  sugar  may  be 
added  for  the  carbon  food. 

Adolf  Mayer's  Culture-Fluid. 
(Unters.  ii.  d.  ale.  Gahr.,  1870.) 

Magnesium  sulphate 10.0 

Ammonium  nitrate 15.0 

Tri-basic  calcium  phosphate .1 

Potassium  phosphate IO.O 

Distilled  water 1,000.0 

Dissolve  cold  and  add  sugar.  Add  sodium 
chloride  (3  per  cent)  if  it  is  to  be  used  for 
luminous  bacteria,  and  an  excess  of  pure  car- 
bonate of  lime  if  acid-forming  bacteria  are  to  be 
grown. 


Uschinsky's  Solution. 

Distilled  water 1,000 

Glycerin    30  to  40 

Sodium  chloride 5  to  7 

Calcium  chloride o.i 

Magnesium  sulphate 0.3  to  0.4 

Dipotassium  phosphate 2  to  2.5 

Ammonium  lactate 6  to  7 

Sodium  asparaginate 3tO4 

Modified  Uschinsky's  Solution. 
The  modified  Uschinsky  recommended  by  the 
writer  for  use  with  starch  jelly  is  made  as  fol- 
lows: 

Distilled  water 1,000.000 

Ammonium  lactate 5.000 

Sodium  asparaginate 2.500 

Sodium  sulphate 2.500 

Sodium  chloride 2.500 

Dipotassium  phosphate 2.500 

Calcium  chloride .010 

Magnesium  sulphate .010 

Fraenkel  and  Voges'  Solution. 
(Hygienische  Rundschau,  Bd.  IV,  1894,  p.  769.) 

Water   1,000 

Sodium  chloride 5 

Dipotassium  phosphate*  2 

Ammonium  lactate 6 

Sodium  asparaginate 4 

This  paper  also  discusses  Uschinsky's  solution. 

Fermi's  Culture-Fluid. 

Distilled  water 1,000.0 

Magnesium  sulphate .2 

Acid  potassium  phosphate i.o 

Ammonium  phosphate 10.0 

Glycerin  45.0 

This  may  be  added  to  agar  in  place  of  pepton- 
ized  beef-broth  (De  Schweinitz)  or  to  silicate 
jelly,  in  which  case  the  volume  of  water  must 
be  reduced  (see  Silicate  Jelly,  p.  36). 

Moore's  Culture-Medium  for  Leguminous  Root- 
tubercle  Bacilli. 
(For  field  use.) 

The  dried  culture  (on  cotton)  is  thrown  into 
clean  water  containing :  Cane-sugar,  I ;  c.  p. 
monopotassium  phosphate,  o.i;  c.  p.  magnesium 
sulphate,  o.oi  per  cent.  After  24  hours  add 
Merck's  pure  dibasic  ammonium  phosphate  to 
amount  of  0.5  per  cent.  Seeds  are  drenched 
with  this  fluid  at  end  of  another  day,  dried  in 
shade,  and  planted. 


*  l.i-i  mi  ami  ami  Neumann  recommend  neutral  commercial  sodium  phosphate  ('.  c.  p.  29.) 


198 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


Maassen's  Culture-Fluid. 

Malic  acid 7 

Distilled  water 100 

Neutralize  to  litmus  exactly  with  7  per  cent 
potassium  hydrate.  Make  up  to  1,000  cc.  with 
distilled  water  and  add : 

Asparagin   10.0 

Secondary  sodium  phosphate 5.0 

Magnesium  sulphate 2.5 

Sodium  hydrate 2.5 

When  dissolved  add  o.oi  gram  of  calcium 
chloride. 

To  this  may  then  be  added  grape-sugar  or  any 
other  carbon  food  desired. 

Proskauer  and  Beck's  Culture-Fluid. 

Distilled  water 1,000.00 

Commercial     ammonium     car- 
bonate           3.50 

Primary  potassium  phosphate..         1.50 

Magnesium  sulphate 2.50 

Glycerin  15.00 

Mackensie's  Culture-Fluid. 

Acid  ammonium  tartrate 

Bipotassium  phosphate 

Potassium  sulphate 

Sodium  chloride 

Glucose  

Lactose  

Glycerin   


1.5- 
2.5 

i.S 
•5 
5-o 
5-0 
15.0 
Water   1,000.0 

This  is  rendered  alkalin  to  phenolphthalein 
with  normal  soda  solution. 

Culture-Medium  for  Luminous  Bacteria. 

(Molisch.,  /.  c.  p.  87.) 

Water   1,000.000 

Gelatin  100.000 

•Sugar 20.000 

Peptone  10.000 

Dipotassium  phosphate .250 

Magnesium  sulphate .250 

Enough  sodium  hydroxid  to  render  the  me- 
dium feebly  alkalin.  On  this  substratum  the 
bacteria  grow  feebly  and  are  not  luminous  until 
sodium  chloride  or  some  equivalent  substance  is 
added  (usually  3  per  cent).  Then  they  grow 
well  and  become  luminous. 


Winogradsky-Sleskin  Silicate  Jelly. 

Ammonium  sulphate 0.40 

Magnesium  sulphate 05 

Potassium  phosphate 10 

Sodium  carbonate 0.60-  .90 

Calcium  chloride Trace 

Silicate  jelly 100.00 

The  salts  are  dissolved  separately  in  the  least 
possible  water,  and  added  to  the  dialyzed  acid. 

(For  further  observations  on  silicate  jelly  see 
P.  36.) 

Nitrogen-free  Medium  for  Bacteria. 

Triple-distilled  water 1,000.000 

Cane-sugar  5.000 

Monopotassium  phosphate. ...          2.000 

Magnesium  sulphate .100 

Sodium  chloride .500 

All  chemically  pure,  in  scrupulously  clean 
flasks.  The  water  should  be  freshly  distilled, 
kept  in  glass-stoppered  bottles,  and  tested  fre- 
quently with  Nessler's  solution  for  presence  of 
ammonia. 

Giltay  &•  Aberson's  Culture-Medium  for  Deni- 
trifying Organisms. 

Distilled   water 1,000.0 

Potassium  nitrate 2.0 

Asparagin   i.o 

Magnesium  sulphate 2.0 

Citric  acid 5.0 

Monopotassium  phosphate 2.0 

'Calcium   chloride .2 

Ferric  chloride 2  gtts. 

The  acid  should  be  neutralized  by  the  addition 
of  potassium  hydrate. 

This  medium  is  a  modification  of  that  of 
Gayon  and  Dupetit,  less  nitrate  being  used  and 
the  neutralization  being  made  with  potash  in- 
stead of  ammonia.  In  preparing  this  fluid  the 
asparagin  and  the  nitrate  of  potash  are  dissolved 
in  250  cc.  of  water;  the  other  substances  are  dis- 
solved in  500  cc.  of  water,  and  after  the  citric 
acid  has  been  neutralized  the  two  fluids  are 
mixed,  cooled  to  15°  C.,  and  sufficient  water 
added  to  make  I  liter.  When  the  nitrate  of 
potash  and  the  asparagin  are  dissolved  along 
with  the  other  salts  a  decomposition  occurs,  and 
the  liquid  is  browned  from  the  presence  of 
nitrous  acid,  which  should  be  avoided.  Some 
carbonate  of  lime  is  also  added  to  the  culture 
fluid.  Instead  of  asparagin,  2  grams  of  dextrose 


SYNTHETIC    CULTURE    MEDIA. 


I99 


may  be  added.  If  the  latter  is  used  the  fluid 
must  not  contain  the  least  excess  of  potassa; 
otherwise  when  it  is  sterilized  there  will  be  more 
or  less  humification.  (Recherches  sur  un  mode 
de  devitrification,  etc.  Archives  neerlandaises 
des  Sci.  Ex.  et  Nat.,  Tome  XXV,  1892,  pp  341- 
36i.) 

Winogradsky's   Culture-Medium   for   Nitrogen- 
Assimilating  Soil-Bacteria. 

Twice-distilled  water 1,000.00 

Potassium  phosphate i.oo 

Magnesium  sulphate 0.50 

Sodium  chloride 01  to  0.02 

Iron  sulphate 01  to   .02 

Manganese  sulphate 01  to   .02 

Dextrose  20      1040 

To  this  should  be  added  a  small  quantity  of 
pure  calcium  carbonate,  30  or  40  grams  per  liter 
is  sufficient.  The  carbonate  is  freshly  washed 
in  boiling  water  and  added  in  paste  or  dried 
rapidly  and  preserved  in  flasks  with  ground- 
glass  stoppers.  It  is  recommended  that  the  sec- 
ond distillation  of  the  water  be  made  with  car- 
bonate of  soda  and  that  pure  salts  be  obtained 
by  repeated  crystallizations.  It  is  probable  that 
monopotassium  phosphate  is  meant  by  phosphat 
de  potasse.  (Recherches  sur  I'assimilation  de 
1'azote  libre  de  1'atmosphere  par  les  microbes, 
Archives  des  Science  Biologiques,  Tome  III, 
p.  304,  St.  Petersburg,  1895.) 

Beyerinck's  Agar  for  Cultivation  of  the  Nitrite 
Bacteria. 

Ordinary  agar  is  added  to  distilled  water, 
heated  until  it  passes  into  solution,  and  poured 
into  Erlenmeyer's  flasks,  where  it  is  left  to 
solidify.  When  cold  the  flasks  are  filled  with 
distilled  water  (not  necessarily  sterile)  and  set 
away.  After  several  changes  of  water  and  the 
lapse  of  one  or  two  weeks  the  soluble  organic 
substances  will  have  been  absorbed  out  of  the 
agar,  and  to  it  may  now  be  added  the  inorganic 
nutrient  substances  desired,  after  which  it  is 
sterilized.  Along  with  the  nutrient  substances 
some  pure  precipitated  calcium  carbonate  should 
be  added.  The  sterile  agar  may  then  be  solidi- 
fied in  Petri  dishes,  test-tubes,  etc.  Beyerinck 
considers  this  medium  better  for  isolation  of  the 
nitrite  ferment  than  the  silicate  jelly.  Hydro- 
gen ammonium  sodium  phosphate  (NHi  Na- 
HPOi-KIW))  is  recommended  as  the  best  one 
of  the  ammonium  salts  for  addition  to  the  agar, 
because,  upon  cooking,  the  agar  is  not  attacked, 
and  consequently  additional  soluble  substances 


are  not  liberated  from  it.  (Beyerinck:  Kultur- 
versuche  mit  Amoben  auf  festem  Substrate. 
Centralb.  f.  Bakt,  i  Abt,  Bd.  XIX,  1896,  pp. 
257-267.) 

Winogradsky's  Nutrient  Agar  for  Isolation  of 
Nitrate  Bacteria. 

Tap-water   I)OOo 

Agar  (thoroughly  washed) 15 

Di-potassium  phosphate 0.05 

Fused  sodium  carbonate i 

Sodium  nitrite  (Merck) 2 

(Centralb.  f.  Bakt.,  2  Abt.,  Bd.  V,  1899,  pp. 
537-549-) 

Winogradsky  &  Omelianski's  Fluid  Culture- 
Medium  for  Isolating  the  Nitrate  Bacteria  from 
Soils. 

Distilled    water. 1,000.0 

Magnesium  sulphate .3 

Ferrous  sulphate .4 

Sodium  chloride .5 

Di-potassium  phosphate .5 

Fused  sodium  carbonate i.o 

Sodium  nitrite  (Merck) i.o 

Transfers  through  a  series  of  flasks  are  neces- 
sary in  order  to  isolate  the  organism.  (Cen- 
tralb. f.  Bakt.,  2  Abt.,  Bd.  V,  1899,  pp.  537-549.) 

Winogradsky  &•  Omelianski's  Fluid  Culture- 
Medium  for  Isolating  the  Nitrite  Bacteria  from 
Soils. 

Distilled  water 1,000.0 

Ferrous  sulphate .4 

Magnesium  sulphate .5 

Di-potassium  phosphate i.o 

Sodium  chloride 2.0 

Ammonium  sulphate 2.0 

The  cultures  are  made  in  broad-bottomed 
flasks  in  50  cubic  centimeters  of  the  fluid,  to 
each  of  which  about  l/2  gram  of  magnesium  car- 
bonate is  added.  It  is  necessary  to  transfer 
through  a  series  of  flasks  in  order  to  obtain 
pure  cultures.  If  the  organism  does  not  grow 
well  on  the  start,  additional  ammonium  sul- 
phate may  be  added,  viz,  i  cubic  centimeter  of  a 
10  per  cent  solution  to  each  flask.  (Centralb. 
f.  Bakt.,  2  Abt,  Bd.  V.,  1899,  pp.  537-549-) 

Dubois'  Fluid  Medium  for  Luminous  Bacteria. 
(See  '93  Dubois,  Bibliog.,  XXVII.) 

Kuntze's  Medium  for  Bacillus  Prodigiosus. 
(See  'oo  Kuntze,  Bibliog,  XXIII.) 


2OO 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Omelianski's  Magnesia-Gypsum  Blocks  for  the 
Cultivation  of  Nitrifying  Organisms. 

One  per  cent  carbonate  of  magnesia  is  uni- 
formly mixed  with  gypsum  and  water  added  to 
it,  stirring  until  it  becomes  of  the  consistency 
of  sour  cream,  when  it  is  poured  upon  plate- 
glass  and  spread  out.  As  soon  as  the  mass  be- 
comes of  a  doughy  consistency  and  is  ready  to 
harden,  it  is  cut  into  circular  blocks  for  Petri 
dishes  and  into  strips  for  test-tubes.  The  cir- 
cular pieces  may  be  cut  with  a  Petri  dish  of  a 
size  a  little  smaller  than  the  dishes  it  is  intended 
to  use.  As  soon  as  the  gypsum  has  hardened 
thoroughly  the  blocks  are  pried  loose  from  the 


plate  glass,  placed  bottom  up  in  the  dishes  (so 
as  to  give  a  smooth  surface),  and  enough  of  the 
above  (nitrite)  culture-medium  added  to  half 
cover  the  block.  This  is  then  autoclaved  and 
additional  sterile  culture-media  added  from 
time  to  time  as  necessary,  being  careful  not  to 
wet  the  inoculated  surface  of  the  block.  The 
sowings  are  made  on  the  smooth  surface  of  the 
block  and  the  dishes  are  kept  in  a  thermostat  at 
25°  to  30°  C.  Colonies  begin  to  be  visible  in 
4  to  5  days.  In  10  to  14  days  many  colonies 
are  0.25  to  0.50  millimeter  in  diameter.  (Cen- 
tralb.  f.  Bakt.,  2  Abt.,  Bd.  V,  1899,  P-  652.) 


MISCELLANEOUS. 


Distilled  Water. 
(See  page  124.) 


Chromic  Acid  Cleaning  Mixture. 

This  is  made  by  pouring  I  gallon  or  more  of 
concentrated  crude  sulphuric  acid  into  an  equal 
volume  of  a  saturated  aqueous  solution  of  po- 
tassium bichromate.  It  should  be  done  in  a 
large  enameled  iron  kettle,  the  acid  being  added 
slowly  at  intervals,  with  frequent  stirring,  so  as 
to  keep  the  mixture  below  the  boiling  point.  An 
excess  of  the  sulphuric  acid  should  be  avoided. 
Pure  water  should  be  used  for  dissolving  the 
potassium  bichromate,  and  under  no  circum- 
stances should  this  solution  be  poured  into  the 
acid,  since  steam  might  be  generated  and  dan- 
gerous splutterings  occur.  The  resulting  chromic 
acid  is  very  injurious  to  the  skin  and  should  be 
used  with  care.  At  15°  C.  each  10  parts  by 
weight  of  water  will  dissolve  about  I  part  of  the 
potassium  salt.  The  chromic  acid  mixture  is 
said  to  explode  violently  when  brought  into  con- 
tact with  certain  substances,  e.  g.,  alcohol,  gly- 
cerin. 

Fluid  for  Softening  Hard  Tissues. 

Frequently  grains  of  cereals  and  other  hard 
tissues  may  be  softened  for  cutting  on  the 
microtome  with  slant  stroke  by  soaking  from 
3  to  6  months  in  equal  parts  of  alcohol  and 
glycerin. 

Unguentum  resinae. 

(See  Bibliog.,  XVII,  'oo,  Bulloch.) 
Darwin's  Wax-Mixture. 

This  consists  of  vaseline  50  parts,  beeswax  35 
parts,  melted  together.  Then  stir  in  of  pow- 
dered resin  15  parts.  If  a  stiffer  mixture  is  de- 


sired, add  more  wax  up  to  50  parts  (see  Darwin 
&  Acton,  Plant  Physiology,  p.  3,  foot  note). 

Pencils  for  Writing  on  Glass. 
(See  page  in.) 

Pyrogallol  Developer. 

(Much  used  in  Laboratory  of  Plant  Pathol- 
ogy.) 

1 i)  Alkali: 
Carbonate     of    potash 

(cryst. )* il/2  ounces,  or  46.5  grams 

Carbonate  of  soda 

(cryst.)* 2  ounces,  or  62  grams 

Distilled  water 12^2  ounces,  or  375  cc. 

(2)  Pyro : 
Sulphite       of        soda 

(cryst.)* 4  ounces,  or  124  grams 

Citric  acid 60  grains,  or  3.9  grams 

Bromide  of  potash..  40  grains,  or  2.6 grams 

Distilled  water 121A  ounces,  or  375  cc. 

Pyrogallic  acid i  ounce,  or  31  grams 

The  pyrogallol  should  be  added  last  of  all,  and 
the  nearly  filled  bottle  closed  at  once. 

For  a  normal  developer  take  2  drams  of  No.  i, 
add  2  drams  of  No.  2,  and  make  up  to  4  ounces 
with  distilled  water.  Reduce  the  amount  of  al- 
kali to  one-fourth  dram  or  less  in  case  of  much 
overexposed  plates.  In  case  of  exposures  likely 
to  exhibit  too  great  contrasts  reduce  the  pyro. 
Always  begin  development  with  one-fourth  of 
the  alkali,  unless  the  exposure  is  known  to  be 
correct.  Both  solutions  should  be  kept  in  glass- 
stoppered  bottles. 

Bottles  containing  alkali  should  have  the  in- 
side of  the  neck  and  the  ground  surface  of  the 
stopper  wiped  dry  before  replacing;  then  the 
latter  will  not  stick. 


•In  case  anhydrous  salts  are  employed,  use  one-half  as  much. 


SYNTHETIC    CULTURE    MEDIA. 


2OI 


Ortol  Developer. 
(See  pp.  140-141.) 

Pyro  Developer  for  Dry  Plates. 
(Recommended  by  S.  G.  Lofft.) 

(1)  Water   iooz.,       or  300      cc. 

Citric  acid 10  grains,  or     6.46  grams 

Pyrogallic  acid...      I  oz.,       or   31      grams 

(2)  Sodium     sulphite 

(crystals)    402.,       or  124      grams 

Water  16  oz.,       or  480      cc. 

Or 

Seed's   sulphite.  ...i 5^  oz.,       or   46.5   grams 
Water   i6oz.,       or  480      cc. 

(3)  Sodium  carbonate 

(crystals)    4oz.,       or  124      grams 

Water   i6oz.,       or  480      cc. 

Or 
Seed's  carbonate..     2oz.,       or  62      grams 

Water  i6oz.,       or  480      cc. 

To  develop  take — 

Water  4  oz.,       or  120      cc. 

No.  i  2  drams,  or     7.5   cc. 

No.  2  J'a  oz.,       or   15      cc. 

No.  3   1A  oz.,       or    15      cc. 

For  underexposures  dilute  and  change  fre- 
quently to  fresh  developer. 

For  overexposures  use  old  developer  or  re- 
strain with  a  few  drops  of  10  per  cent  potassium 
bromide. 

Lantern-slide  Developer. 

(Used  in  Laboratory  of  Plant  Pathology.) 

Distilled  water cc. .       900 

Carbonate  of  soda  (cryst.)  grams. .     39 
Sulphite  of  soda  (cryst.).  ..grams. .     39 

Hydrochinon   grams..     13 

Add  the  hydrochinon  after  solution  of  the 
soda  salts,  and  put  at  once  into  a  glass-stoppered 
bottle.  For  use  take  3  ounces  of  above  and  3 
ounces  of  distilled  water,  to  which  add  5  drops 
of  a  10  per  cent  solution  of  bromide  of  potas- 
sium. If  properly  exposed  the  image  should 
appear  in  30  to  60  seconds,  and  the  development 
should  be  over  in  3  or  4  minutes.  Good  for  a 
dozen  or  more  properly  exposed  slides. 
Zettnow's  Copper-chrom-filter. 

Dry,  pure,  copper  nitrate 160 

Pure  chromic  acid 14 

Distilled  water 250 

This  may  be  diluted  further  with  water  if 
desired. 

In  case  there  is  difficulty  in  preparing  the 
above,  a  solution,  stated  by  Zettnow  to  be  nearly 
as  useful,  consists  of — 


Sulphate  of  copper 175 

Bichromate  of  potash 17 

Water  1,000 

(Centralb.  f.  Bakt.,  IV  Bd.,  1888,  p.  51.) 
Neuhaus  says,  water  500  to   1,000,  and  also 
2  cubic  centimeters  of  sulphuric  acid.    This  so- 
lution extinguishes  all  the  blue  and  violet  rays. 

Toning  Bath  for  Solio  Paper. 

(A)  Hyposulphite  of  soda. ..     8 ounces 
Potash  alum  (crystals)..    6 ounces 

Cane  sugar  2  ounces 

Water 80  ounces 

Dissolve  cold,  then  add  2  ounces  of  borax 
which  has  been  dissolved  in  8  oz.  of  hot  water. 
Let  stand  12  hours,  and  then  decant. 

(B)  Pure  chloride  of  gold. .     7.5  grains 

Acetate  of  lead 64    grains 

Distilled  water 8  ounces 

This  solution  must  not  be  filtered  and  must 
be  shaken  thoroughly  each  time  before  using. 
Solio  paper  should  be  printed  about  one-third 
darker  than  it  is  desired  to  have  the  pictures. 
When  the  prints  are  ready  they  are  placed  face 
down  in  a  toning  mixture  made  of  stock  A,  8 
ounces,  and  stock  B,  i  ounce,  taking  care  that 
the  entire  surface  of  each  print  is  uniformly 
wetted.  They  are  allowed  to  remain  in  this 
bath,  with  constant  movement  by  means  of  the 
fingers,  until  the  desired  color  is  obtained  (usu- 
ally about  5  minutes).  The  prints  are  now  im- 
mersed in  salt  water  (i  -.32)  for  5  minutes. 
They  are  then  exposed  for  15  minutes  to  the 
fixing  bath,  consisting  of — 

Hyposulphite  of  soda I     ounce 

Sulphite  of  soda  (crystals)  60     grains 

Borax  1A  ounce 

Water  20     ounces 

The  prints  are  finally  washed  for  from  i  to  2 
hours  in  running  cold  water.  The  toning  bath 
should  not  be  cooler  than  40°  or  warmer  than 
60°  F. 

A  New  Test  for  Indol. 

Herter  &  Foster  have  recently  described  what 
is  stated  to  be  a  rapid  and  accurate  method  of 
determining  indol,  adapted  either  for  colorimet- 
ric  or  gravimetric  determinations.  To  slightly 
alkaline  solutions  naphthoquinon  sodium  mono- 
sulfonate  is  added.  A  blue  crystalline  com- 
pound, di-indyl  naphtho-ketone  mono-sulfonate 
is  produced.  This  is  slightly  soluble  in  water, 
but  is  readily  soluble  in  chloroform,  its  solution 
being  red.  (Science,  n.  s.,  Vol.  XXI,  1905, 
p.  987.) 


2O2 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


FIXING  FLUIDS. 


Absolute  Alcohol. 

Expose  24  hours  or  more.  Very  useful  for 
fixing  bacteria  in  tissues,  as  it  prevents  their 
diffusion.  It  causes,  however,  considerable 
shrinkage  of  the  tissues,  and  the  nuclei  are  often 
difficult  to  stain,  and  are  usually  distorted. 
Bacteria  fixed  in  this  way  stain  well  in  Ziehl's 
carbol-fuchsin. 

Picric  Acid  in  Hot  Absolute  Alcohol. 
(See  p.  8.) 

Mercuric  Chloride  in  Hot  Absolute  Alcohol. 
(See  p.  8.) 
Acetic  Alcohol  ivith  Mercuric  Chloride. 

Absolute  alcohol I 

Glacial  acetic  acid i 

Chloroform  i 

Add  mercuric  chloride  until  saturated.  Wash 
with  alcohol  or  with  alcohol  containing  tincture 
of  iodine.  One  of  the  most  rapid  fixatives 
known. 

Acetic  Alcohol  (Camay's  Fluid.} 

Glacial  acetic  acid. i 

Absolute  alcohol ' 3 

Expose  24  hours  or  more  and  wash  in  alcohol. 
This  solution  preserves  the  chromatic  and  cyto- 


plasmic  structures  better  than  alcohol  alone,  and 
shrinks  the  tissues  much  less.  Tissues  fixed  in 
this  solution  take  most  of  the  coal-tar  stains 
better  also.  It  has  very  great  penetrating  power, 
which  makes  it  very  useful  in  fixing  large  pieces 
of  tissue. 

Chromo-aceto-osmic  Acid  (Flemming's  Fluids). 

A.  Stronger  solution — • 

1  per  cent  chromic  acid IS 

2  per  cent  osmic  acid 4 

Glacial  acetic  acid i 

B.  Weaker  solution — • 

i  per  cent  chromic  acid 25 

I  per  cent  osmic  acid 10 

i  per  cent  acetic  acid 10 

Water  55 

Expose  12  to  48  hours  and  wash  in  running 
water.  This  is  probably  the  most  valuable  of 
all  the  fixing  fluids  for  preserving  exactly  all  the 
cell  structures;  and  material  fixed  in  it  takes  the 
coal-tar  stains  unusually  well.  It  has  very 
slight  penetration,  and  therefore  the  tissues 
should  be  cut  into  pieces  not  more  than  2  milli- 
meters thick.  It  should  be  kept  in  glass-stop- 
pered bottles,  and  should  be  made  fresh  every 
6  months  or  less. 


For  additional  formula;  consult  the  various  standard  text-books,  especially  Eyre's 
"  Bacteriological  Technique,"  where  may  be  found,  among  others,  the  following: 


Kitasato's  glucose-formate-bouillon . 
Iron-bouillon.         L,ead-bouillon. 
Parietti's  bouillon. 
Carbolized  bouillon. 
Kitasato's  glucose-formate-agar. 
Guarniari's  agar-gelatin. 
Carbolized  agar. 
Glycerine  blood -serum. 
Heiman's  serum-agar. 
Washbourn's  blood-agar. 
Urine- gelatin.          Urine-agar. 
Whey-agar.  Fish-bouillon. 

Fish-gelatin.  Fish-agar. 


Glycerinated  potato. 
Glycerine-potato-broth . 
Eisner's  potato-gelatin. 
Goadby's  potato-gelatin. 
Beer- wort.         Wort-gelatin. 
Wort-agar.        Wine-must. 
Gasperini's  wheat-broth. 
Bread-paste.         Milk-rice. 
Fakes'  iron-peptone-solution. 
MacConkey's  bile-salt-broth. 
MacConkey's  bile-salt-agar. 
Sabouraud's  French  proof  agar. 
Blaxall's  English  proof  agar. 


PLATE  24. 


Brown  rot  of  the  potato. 

Potato-tuber  from  Hastings,  Florida,  crop  of  1905.  sound  externally,  but  brown-rotted  in  the  vascular  system,  the  bacteria  having  entered 
through  the  vessels  of  the  underground  stem.  Bacterium  solanacearum  was  plated  out  and  subculture*  were  used  to  infect  potted  potato- 
plants.  The  plant  shown  was  inoculated  on  June  27,  by  delicate  needle- pricks.  Disease  more  advanced  on  one  inoculated  «hoot  than  on 
the  other;  later  this  also  shriveled.  Tuber  natural  size;  photographed  May  29.  Plant  about  one-fifth  natural  size;  photographed 
July  15.  1905. 


PLATE  25. 


Early  Rose  potatoes  destroyed  by  Bacterium  solanacearum. 

Plants  inoculated  at  same  time,  in  same  way.  and  from  same  source  as  plant  shown  in  plate  24.     About  one-fourth  natural  size.     Progress  of  disease  rather 
slow  ;  no  tubers  were  formed.     All  of  the  shoots  dead  or  dying.     Photographed  August  3,  1905,  i.  e.  at  end  of  fifth  week. 


PLATE  26. 


Effect  of  Bacterium  solanacearum  on  a  tomato-plant. 

The  left-hand  plant  was  inoculated  in  the  stem  on  July  5,  by  means  of  a  few  needle-pricks.  The  right-hand  plant  was  pricked  with  a  sterile  needle. 
The  infectious  material  was  a  subculture  from  a  poured-plate  colony.  This  was  obtained  from  the  interior  of  a  potato-stem,  naturally  infected,  in 
District  of  Columbia.  Photographed  July  I  1 .  1 904.  About  one-third  natural  size. 


PLATE  27. 


The  Granville  tobacco-wilt,  a  bacterial  disease. 

The  left-hand  plant  wilted  naturally  in  the  field,  was  pruned,  transplanted  to  the  hothouse,  recovered  for  a  few  weeks,  developed  the  leaves  shown,  and  then  wilted  again. 
Photographed  June  28;  one-thitd  natural  size. 

In  this  disease  the  vascular  ring  is  browned  and  cavities  are  formed  in  the  stem.  The  bacterial  slime  is  gray-white  and  abundant.  Fungi  were  not  present  in  the  plants 
examined  hy  me.  Poured  plates  were  made  from  the  interior  of  several  such  stems,  and  these  yielded  practically  pure  cultures  of  one  organism.  A  subculture  from  one 
of  these  colonies  was  used  to  inoculate  the  right-hand  plant.  The  needle-pricks  were  made  on  July  13,  1905.  Photographed  August  3;  one-half  natural  size.  Twelve 
plants  were  inoculated  at  this  time  and  all  contracted  the  disease.  The  signs  and  lesions  were  the  same  as  in  the  plants  obtained  from  the  field.  The  organism  causing 
this  disease  is  closely  related  to  Bact.  solanacearum. 


PLATE  28. 


Pear-shoots  blighted  by  Bacillus  amylovorus. 

Inoculation*  by  needle-priclcson  rapidly  growing  items.  The  infectious  material  came  from  a  green  apple.  The  poured  plates  yielded  practically  pure  cultures  and 
the  inoculations  were  made  directly  from  colonies  in  these  plates.  Of  1 2  inoculations  (8  shoots,  4  fruits)  II  were  successful.  Needle-pricks  made  July  10,  1905. 
Photographed  July  20.  The  lower  leaves  were  still  green  ;  the  tops  (A  and  A)  had  shriveled  and  browned  ;  the  bacteria  had  passed  downward  in  the  bark  to 
points  below  B  and  B,  and  they  had  also  run  out  into  the  petioles  (P  and  P)  and  had  browned  them,  but  the  blades  of  these  leaves  were  still  green. 


PLATE  29. 


Green  pears  inoculated  with  Bacillus  amylovorus. 

These  were  inoculated  from  the  same  poured  plate  and  at  the  same  time  as  the  shoots  shown  in  plate  28,  but  the  photographs  were  made 
two  days  earlier.  The  inoculations  were  by  means  of  a  few  needle- punctures.  The  inoculated  parts  browned,  softened,  shriveled 
slightly,  and  were  extruding  bacterial  slime  from  small  cracks  and  from  many  stomata.  Beyond  the  browned  area  there  was  a  water- 
soaked  area.  The  internal  injury  was  extensive — m  one  fruit  nearly  the  whole  interior  had  softened  and  was  occupied  by  the 
bacteria.  These  tissues  were  filled  with  grayish  bacterial  slime  to  such  an  enormous  extent  that  on  handling  them  the  fingers 
dripped  with  it.  Inoculated  July  10,  1905.  Photographed  July  18;  about  natural  size.  Poured  plates  made  from  the  interior  of 
these  fruits  yielded  pure  cultures  of  B.  amylovorus. 


PLATE  30. 


Blighted  quince-shoots  and  pear-fruits. 

Twelve  inoculations  were  made  and  all  were  successful — 8  quince-shoots  and  4  pear-fruits.  None  of  the  checks  contracted  the  disease.  The  organism  was 
introduced  by  delicate  needle- pricks.  The  subculture  used  was  derived  from  a  poured-plate  colony  obtained  from  the  interior  of  one  of  the  pears  shown  in 
plate  29.  Owing,  no  doubt,  to  the  riper  and  therefore  less  susceptible  condition  of  the  tissues,  and  possibly  also  to  somewhat  cooler  weather,  the  disintegration 
of  the  deeper  tissues  of  the  fruits  had  not  proceeded  as  far  as  in  the  pears  inoculated  on  the  same  tree  1 0  days  earlier ;  in  those  fruits  first  inoculated  the  decay 
was  from  5  to  10  times  as  extensive  on  the  8th  day  as  in  the  second  inoculations  on  the  11th  day.  Inoculated  July  20,  1905.  Photographed  July  31. 


PLATE  31. 


Small  green  apples  blighted  by  Bacillus  amylovorus. 

One  of  these  fruits  furnished  the  bacteria  used  to  inoculate  the  shoots  and  fruits  shown  in  plates  28  and  29.  The  interior  of  these  fruits  was  gorged 
with  a  gray-white  bacterial  slime.  The  surface  was  browned  and  shriveling  in  many  placet  and  water-soaked  in  others.  Bacteria  were  oozing 
from  the  darker  parts.  Every  one  of  many  fruits  examined  had  been  wounded  by  the  curculio,  and  probably  infection  occurred  in  this  manner. 
Arlington  Farm,  Virginia.  Photographed  June  30,  1905. 


BIBLIOGRAPHY. 


(jeneral  Literature. 


As  far  as  possible  the  references  in  this  bibliography  have  been  arranged  chronologically  so  that  the 
reader  may  see  at  a  glance  the  historic  development  of  the  subject.  In  several  cases  logic  has  been  sacrificed 
to  convenience,  e.  g.,  in  keeping  the  nitrifying  and  denitrifying  bacteria  separate  from  the  group  devoted  to 
oxidation  and  reduction,  the  average  reader,  it  was  thought,  being  more  likely  to  search  for  references  to  the 
nitrogen  bacteria  under  the  former  than  under  the  latter  heading. 


I.    Journals. 

Archiv  fiir  Hygiene.  Munich  and  Leipsic.  Verlag 
von  R.  Oldenbourg. 

Begun  in  1883.    51  vols.  to  date. 

Zcitschrift  fur  wissenschaftliche  Mikroskopie  und 
fiir  mikroskopische  Technik.  Braunschweig. 
Harald  Bruhn. 


Begun  in  18 


21  vols.  to  date. 


Zeitschrift  fur  Hygiene  und  Infectionskrankheiten. 
Leipsic.  Veit  und  Comp. 

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Annales  de  1'Institut  Pasteur.  Paris,  G.  Masson 
et  Cie. 

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Begun  in  1887  under  the  title,  Centralb.  f.  Bakt.,  und 
Parasitenkunde.  Title  changed  and  scope  limited  when 
the  following  publication  was  begun.  37  vols.  to  date. 

Central'blatt  fiir  Bakteriologie,  Parasitenkunde  und 
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Annales  de  Micrographie,  specialement  consacrees  a 
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Begun  in  1888. 

Zeitschrift  fiir  Pflanzenkrankheiten.  Paul  Sorauer, 
editor.  Verlag  von  Eugen  Ulmer,  Stuttgart. 


Begun  in  1891. 
only  on  bacteria. 


n  vols.   to  date.     Occasional  articles 


The  Journal  of  Pathology  and  Bacteriology.     Edin- 
burgh and  London.    Young  J.  Pentland. 

Begun  in  1892.    9  voU.  to  date. 

Journal  of  the  American  Public  Health  Association. 
The  number  for  October,  1895,  contains  numerous  valu- 
able papers  on  bacteriology.  The  number  for  October, 
1897,  contains  report  of  a  committee  on  the  pollution  of 
water  supplies;  also  reportof  committee  on  disinfectants. 
Report  of  meeting  held  October  3i-November  3,  1899.  con- 
tains notes  on  the  reaction  at  which  the  maximum 
growth  of  bacteria  occurs  in  liquid  media  ;  a  paper  on 
the  making  of  nutrient  agar-agar  ;  a  paper  on  the  classi- 
fication of  water  bacteria,  etc. 


Tijdschrift  over  Plantenziekten.    Onder  redactie  van 
Ritzema  Bos  en  G.  Staes.    Gent.  J.  Vuylsteke. 

Begun  in  1895.  7  parts  have  appeared.  Only  occasional 
papers  on  bacteria. 

The  Journal  of  Experimental  Medicine.    New  York, 
D.  Apple/ton  &  Co. 

Beguu  in  1896.  Edited  by  Wm.  H.  Welch,  of  Johns 
Hopkins  University.  6  vols.  to  date.  Contains  many 
papers  of  interest  to  persons  who  are  not  physicians. 
Recently  transferred  to  the  Rockefeller  Institute  under 
editorship  of  Simon  Flexner. 

Archives  de  Parasitologie.     Paris,  Geo.  Carre  et  C. 
Naud. 


Begun  in  i& 
by  Societe  d't 


1.    8  vols.  to  date.     Published  since  vol.  4 
itioiis  scientifiques. 


Journal  of  Applied  Microscopy.  Rochester,  N.  Y., 
Bausch  and  Lomb. 

Begun  in  1898.    6  vols.    Discontinued. 

Archives  des  Sciences  Biologiques.  Publiees  par 
1'Institut  imperial  de  medecine  experimentale  a 
St.  Petersbourg. 

Valuable.    Begun  in  1892.     10  volumes  to  date. 

The  Botanical  Gazette.    University  of  Chicago  Press. 

Begun  privately  in  1875.  38  vols.  to  date.  Papers  on 
bacteria  only  in  recent  volumes. 

Journal  of  Hygiene,  Cambridge,  England.  Edited 
by  Nuttall. 

Begun  in  1901.  Published  quarterly  by  Cambridge  Uni- 
versity Press,  Warehouse  Ave.,  Maria  i.;uu-,  London,  E.  C. 
4  vols.  to  date. 

Journal  of  Medical  Research.  Edited  by  Harold  C. 
Ernst,  Boston,  Mass. 

7  vols.  to  date.  This  is  a  continuation  of  the  Journal  of 
the  Boston  Society  of  Medical  Sciences. 

Zeitschrift  fiir  Physikalische  Chemie,   Stochiometrie 
und    Verwandtschaftslehre.      Herausgegeben    von 
Wilh.   Ostwald  und  J.   H.   Van't   Hoff,   Leipzig. 
Verlag  von  Wilhelm  Engelmann. 
About  45  vols.  to  date. 

The  Journal  of  Infectious  Diseases.  Founded  by  the 
Memorial  Institute  for  Infectious  diseases,  Chi- 
cago. 

Begun  in  1904. 

203 


204 


BACTERIA    IN    RELATION   TO    PLANT   DISEASES. 


Bulletin  de  1'Institut  Pasteur.  Revues  et  analyses 
des  travaux  de  microbiologie,  medicine,  biologic 
generate,  physiologic,  chimie  biologique  dans  leurs 
rapports  avec  la  bacteriologie.  Comite  de  redac- 
tion: G.  Bertrand,  A.  Besredka,  A.  Borrel,  C.  Dele- 
zenne,  A.  Marie,  F.  Mesnil,  dp  1'Institut  Pasteur 
de  Paris. 

Begun  in  1903. 

II.    Transactions,  Beitrage,  Jahresberichten, 

Festschriften,  Etc. 

('7°-'92)-  Beitrage  zur  Biologic  der  Pflanzen.    Edited 
by  Ferdinand  Cohn.     15  Heften.     (5  vols.) 
Breslau,  1870-1892. 

('86).  Jahresbericht  iiber  die  Fortschritte  in  der 
Lehre  von  den  Pathogenen  Mikroorganis- 
men  umfassend  Bakterien,  Pilze  und  Pro- 
tozoen.  Von  P.  Baumgarten.  Braun- 
schweig, Harald  Bruhn,  and  more  recently 
Leipzig,  S.  Hirzel. 

Begun  in  1886.  First  volume  covers  the  year  1885.  The 
last  volume  (iSJahrgang)  brings  the  literature  refer- 
ences down  to  the  close  of  1902.  Contains  nothing  on 
plant  parasites,  and  the  title  is  therefore  misleading. 

('86).  Arbeiten  aus  dem  Kaiserlichen  Gesundheits- 
amte.  Berlin,  Verlag  von  Julius  Springer. 

Extremely  valuable  on  account  of  the  high  quality  of 
the  papers,  which  deal  mostly  with  animal  diseases.  The 
first  two  volumes  were  published  as  Mitteilungen,  etc. 
Begun  in  1886. 

('92-'95).  Beitrage  zur  Physiologic  und  Morphologic 
niederer  Organismen.  W.  Zopf.  5  Heften. 
Leipsic,  Arthur  Felix,  1892-1895. 

('93).  The  Wilder   Quarter-Century   Book.     Ithaca, 

N.  Y.,  Comstock  Pub.  Co.,  1893. 
Contains  several  valuable  papers. 

Publications  of  the  Division  of  Vegetable  Physiology 
and  Pathology,  Bureau  of  Plant  Industry, 
U.  S.  Department  of  Agriculture,  Washing- 
ton, D.  C. 

Frequently  when  editions  are  exhausted, copies  may  be 
procured  at  actual  cost  of  printing  by  addressing  the 
Bureau  of  Documents,  Union  Building,  Washington, 
D.  C.  The  publications  of  the  Department  of  Agricul- 
ture are  listed  each  mouth,  and  this  list  may  be  obtained 
gratis  on  application  to  the  Secretary  of  Agriculture. 

('94).  KLEIN,  L.,  UND  MIGULA,  W.  Arbeiten  aus  dem 
bacteriologischen  Insititut  der  technischen 
Hochschule  zu  Karlsruhe.  Bd.  I,  n,  in, 
Wiesbaden,  O.  Nemnich,  1894-1903. 

('03).  Contributions  to  medical  research  dedicated  to 
Victor  Clarence  Vaughan  by  colleagues  and 
former  students  of  the  department  of  medi- 
cine and  surgery  of  the  University  of  Michi- 
gan on  the  twenty-fifth  anniversary  of  his 
doctorate.  Ann  Arbor,  George  Wahr,  1903, 
pp.  vi,  620,  with  numerous  plates  and  figures. 


III.    Manuals. 

('80).  WINTER,  GEORG.  Bacteria.  In  Die  Pilze  of 
Rabenhorst's  Kryptogamenflora  Deutsch- 
lands.  I  Abt,  1884. 

The  volume  was  published  in  parts,  the  one  on  bacteria 
coming  out  at  an  early  date  ( 1880) .  Transl.  by  T.  J.  Burrill , 
as  part  of  a  paper  entitled,  "  The  Bacteria,"  Springfield, 
111.,  1882.  nth  Rep.  111.  State  Industrial  Univ. 

('83).  DUCLAUX,  E.  Chimie  biologique — Micro- 
biologie. Paris,  1883,  pp.  vn,  908.  This 
forms  Tome  ix,  first  section,  of  Frem/s  En- 
cyclopedic Chimique. 


('84).  DE  BARY,  ANTON.  Vergleichende  Morphologic 
und  Biologic  der  Pilze  Mycetozoen  und  Bac- 
terien.  Leipzig,  Engelmann,  1884,  pp.  xvi, 
558. 

English  translation  by  Garusey  revised  by  Balfour  in 
1887  under  title  of  "  Comparative  morphology  and  biol- 
ogy of  the  fungi  mycetozoa  and  bacteria."  Oxford,  Clar- 
endon Press. 

('85).  DE  BARY,  ANTON.    Vorlesungen  iiber  Bacte- 

rien.     Leipzig,  Engelmann,  1885,  pp.  vi,  146. 

There  is  an  English  translation  by  Garnsey,  revised  by 

Balfour,  Oxford,  1887.    The  second  German  edition  was 

published  in    1887,   pp.  vi,  158,  20  figs.     A   third  edition 

revised  by  Misrula,  containing  186  pages  and  41  figures, 

appeared  in  1900. 

('86?).  SCHROETER,  J.     Schizomycetes.     In  Die  Pilze 
Schlesiens,    pp.     136-174.      Breslau,    J.    U. 
Kern's  Verlag. 
The  volume  as  a  whole  bears  date  of  1889. 

('87).  LOEFFLER,  FRIEDRICH.  Vorlesungen  iiber  die 
geschichtliche  Entwickelung  der  Lehre  von 
den  Baoterien.  Erster  Theil  bis  zum  Jahre, 
1878,  pp.  xii,  252,  with  3  plates  and  37  text 
figures.  Leipsic,  F.  C.  W.  Vogel,  1887. 

('90).  BAUMGARTEN,  P.  Lehrbuch  der  pathologischen 
Mykologie,  pp.  ix,  973,  24.  Braunschweig, 
Harald  Bruhn,  1890. 

('90).  CORNIL,  A.  V.,  ET  BABES,  V.  Les  bacteries  et 
leur  role  dans  1'etiologie,  Tanatomie  et  Fhis- 
tologie  pathologique  des  maladies  infec- 
tieuses.  3d  ed.  2  vols.  Tome  I,  pp.  vn,  582. 
Tome  n,  pp.  608.  Paris,  Felix  Alcan,  1890. 

('90).  FRAENKEL,  CARL.  Grundriss  der  Bakterien- 
kunde.  3d  ed.  pp.  vin,  515.  Berlin,  August 
Hirschwald,  1890. 

An  excellent  book,  but  now  requires  revision.  There 
is  an  English  translation  by  Linsley.  New  York  :  Wm. 
Wood  &  Co.,  1891. 

('90).  KRAMER,  ERNST.  Die  Bakteriologie  in  ihren 
Beziehungen  zur  Landwirtschait  und  den 
landwirtsdhaftJich  -  technischen  Gewerben. 
Theil  i.  Die  in  der  Landwirtschaft  durch 
Bakterien  bewirkten  Vorgange.  Mit  36 
Abbild.  Wien,  Verlag  von  Karl  Gerolds 
Sohn,  1890.  8vo.  171  pp.  n  Theil.  Die 
Bakterien  in  ihren  Verhaltnisse  zu  den  land- 
wirtschaftlichen-technischen  Gewerben.  8vo. 
pp.  vi,  178,  with  79  figures.  Vienna,  Gerolds 
Sohn,  1891. 

('91).  EISENBERG,  JAMES.  Bakteriologische  Diagnos- 
tik.  3d  ed.  pp.  xxxi,  509.  Hamburg  and 
Leipsic,  Leopold  Voss,  1891. 

('91).  HUEPPE,     FERDINAND.       Die     Methoden     der 
Bakterien-Forschung.      Handbuch    der    ge- 
sammten    Methoden    der    Mikrobiologie.     5 
verbesserte  Aufl.    8vo.    pp.  vm,  495,  with  2 
lithographic  plates  and  68  wood  cuts.   Wies- 
baden, C.  W.  Kreidel's  Verlag,  1891. 
A  French   translation  of  an    earlier  edition,  by  Van 
Ermengem,  entitled,  Manuel  technique  de  microbiolo- 
gie, was  published  at  Paris  in  1887. 

('91).  WOODHEAD,  GERMAN  SIMS.  Bacteria  and  their 
products.  London,  Walter  Scott,  Ltd.,  1891. 
I2mo.  pp.  xin,  459,  with  20  photomicro- 
graphs. 

('92).  STERNBERG,  GEO.  M.  Manual  of  Bacteriology, 
pp.  xn,  886.  8  plates  and  268  figures  in  the 
text.  8vo.  New  York,  Wm.  Wood  &  Co., 
1892. 


MANUALS. 


205 


('94).  FRANKLAND,  PERCY,  AND  FRANKLAND,  MRS.  G. 
C.  Micro-organisms  in  water.  London, 
Longmans,  Green  &  Co.  pp.  xi,  532,  with 
figures,  1894. 

('95).  ABBOTT,  A.  C.  The  principles  of  bacteriology. 
A  practical  manual  for  students  and  physi- 
cians. 3d  ed.  pp.  xii,  493,  with  98  figures, 
17  of  which  are  colored.  Lea  Brothers  & 
Co.,  Philadelphia,  1895.  Last  (6th)  ed.  in 
1902,  pp.  xi,  641. 

('95).  ITZEROTT  UND  NIEMANN.  Mikrophotographi- 
scher  Atlas  der  Bakterienkunde.  Mit  126 
mikrophotographischen  Abbildungen  in 
Lichtdruck  auf  21  Tafeln.  Leipzig,  Johann 
Ambrosius  Barth  (Arthur  Meiner),  1895. 

('95).  KANTHACK  AND  DRYSDALE.  Elementary  prac- 
tical bacteriology,  pp.  xxu,  181,  with  16 
figures,  izmo.  London  and  New  York, 
Macmillan  &  Co.,  1895. 

('95).  FRAENKEL,  CARL  UND  PFEIFFER,  RICHARD. 
Mikrophotographischer  Atlas  der  Bakterien- 
kunde. 2d  ed.  Berlin,  1895.  Verlag  von 
Aug.  Hirschwald.  76  plates  (156  figures) 
with  descriptive  text.  8vo. 

('96).  HUEPPE,  FERDINAND.  Naturwissenschaftliche 
Einfiihrung  in  die  Bakteriologie.  pp.  vm, 
268,  with  28  wood  cuts.  Wiesbaden,  C.  W. 
Kreidel's  Verlag,  1896. 

English  translation  by  Edwin  O.  Jordan,  entitled,  The 
Principles  of  Bacteriology.  The  Open  Court  Pub.  Co  , 
Chicago,  1899. 

('96).  LEHMANN,  K.  B.,  AND  NEUMANN,  RUDOLPH. 
Bakteriologische  Diagnostik.  2  vols.  I2mo. 
Munohen,  J.  F.  Lehmann,  1896.  Theil  I. 
Atlas  of  63  colored  plates.  Theil  n,  Text, 
which  also  includes  some  figures,  pp.  vn, 
448,  and  a  folded  sheet,  giving  a  synopsis  of 
some  of  the  characters  of  62  species. 

There  is  an  English  edition  in  one  volume,  issued  by 
Wm  Wood  &  Co.  in  1897.  The  German  edition  is  a  very 
useful  book.  The  English  edition  does  not  give  the 
translator's  name.  It  also  omits  the  extended  classifica- 
tion and  description  of  species  aud  the  folded  sheet. 

('96).  FLUEGGE,  C.  Die  Mikroorganismen.  Mit  be- 
sonderer  Beriicksichtigung  der  Aetiologie 
der  Infektionskrankheiten.  3  vollig  um- 
gearbeitete  Aufl.  8vo.  2  vols.  Bd.  I.  pp. 
xvi,  596,  with  57  text  figures.  Bd.  u.  pp. 
xxn,  751,  with  153  text  figures.  Leipsic, 
F.  C.  W.  Vogel,  1896. 
Very  useful. 

('96).  STERNBERG,  GEO.  M.    Text-book  of  bacteriol- 
ogy-   PP-  xi,  693,  with  9  heliotype  and  chro- 
molithograph   plates    and    200    engravings. 
8vo.    New  York,  Wm.  Wood  &  Co.,  1896. 
In  the  second  revised  edition  (1901)  there  is  a  chapter 
on  plant  parasites,  but  the  saprophytes  are  omitted  alto- 
gether. 

('96).  CROOKSHANK,  EDGAR  M.  Bacteriology  and  in- 
fective diseases.  4th  ed.  pp.  xxx,  715,  with 
22  plates  and  273  wood  engravings  and  pho- 
tographs. Small  8vo.  London,  H.  K. 
Lewis,  1896. 

('97).  MAC£,  E.  Traite  pratique  de  bacteriologie.  30 
ed.  pp.  vm,  1144,  with  figures.  Paris,  J.  B. 
Bailliere  et  fils.,  1897. 

4th  ed.,  Paris,  1901,  pp.  vm,  1196,  338  figs.  Much  new 
matter  added.  A  very  useful  book. 

('97).  MALI.ORY,  FRANK  BURR,  AND  WRIGHT,  JAMES 
HOMER.  Pathological  technique.  A  prac- 
tical manual  for  the  pathological  laboratory. 


PP-  397,  with  105  illustrations.    Philadelphia, 
W.  B.  Saunders,  1897.    2d  ed.   (with  slight 
change  of  title),  revised  and  enlarged.    1901, 
pp.  432,  with  137  illustrations. 
Compact  and  useful. 

('97).  MIGULA,  W.  System  der  Bakterien.  Hand- 
budi  der  Morphologic,  Entwickelungsge- 
schichte  und  Systematik  der  Bakterien.  i 
Bd.  Allgemeiner  Teil.  8vo.  pp.  vm,  368, 
with  six  plates,  five  of  them  from  photo- 
micrographs. Jena,  Gustav  Fischer,  1897. 

('97).  FISCHER,  ALFRED.  Vorlesungen  iiber  Bakterien. 
8vo.  pp.  vm,  186,  with  28  figures.  Jena, 
Gustav  Fischer,  1897. 

For  criticisms  on  that  portion  of  this  book  which  re- 
lates to  plant  diseases,  see  Centralb.  f  Bakt.,  i  Abt.  1899 
p.  271,  809,  and  1901,88,128,  190.  English  translation  in 
1900  by  A.  Coppen-Jones,  under  title,  The  structure  and 
functions  of  bacteria.  2  German  ed.  in  1903. 

('97-'o3).  LAFAR,  FRANZ.  Technische  Mykologie. 
Ein  Handbuch  der  Garungsphysiologie  fur 
technische  Chemiker,  Nahrungsmittel- 
Chemiker,  Gahrungstechniker,  Agrikultur- 
chemiker,  Pharmaceuten  und  Landwirte. 
Jena,  Gustav  Fischer,  1897-1903.  i  lith.  and 
158  text  figs. 

The  first  volume,  devoted  to  the  bacteria,  contains 
pp.  xii,  362.  This  was  translated  by  Salter  as  Technical 
Mycology,  etc.  London  :  Chas.  Griffin  &  Co.,  and  Phila- 
delphia :  Lippincott,  1898.  A  new  ed.  is  being  published. 

('98).  DUCLAUX,  E.  Traite  de  Microbiologie.  T.  i, 
Microbiologie  general.  Paris,  Masson  et  Cie. 
1898.  pp.  in,  632,  61  text  figures. 

('98).  Procedures  recommended  for  the  study  of 
bacteria,  with  especial  reference  to  greater 
uniformity  in  the  description  and  differen- 
tiation of  species.  Being  a  report  of  a  com- 
mittee of  American  bacteriologists  to  the 
Committee  on  the  Pollution  of  Water  Sup- 
plies of  the  American  Public  Health  Asso- 
ciation. (Submitted  at  the  meeting  of  the 
association  in  Philadelphia,  Sept.,  1897.) 
Concord,  N.  H.,  The  Rum  ford  Press,  1898. 
pp.  47,  with  charts. 

This  pamphlet  should  be  in  every  laboratory,  and  in 
the  hands  of  every  student. 

('98).  HEIM,  L.  Lehrbuch  der  Bakteriologie  mit 
bespnderer  Beriicksichtigung  der  bakterio- 
logischen  Unitersuchung  und  Diagnostik.  2 
Aufl.,  pp.  xvn,  604.  Stuttgart,  Ferdinand 
Enke,  1898. 

('98).  PEARMAIN,  T.  H.,  AND  MOOR,  C.  G..  Applied 
bacteriology,  pp.  xv,  464,  with  text  figures 
and  8  lithographic  plates  (the  latter  copied 
from  Lehmann  and  Neumann),  also  folded 
plate,  after  Stoddard,  showing  3  "halo  cul- 
tures." 2d  ed.,  1898.  London,  Bailliere, 
Tindall  &  Cox. 

('oo).  MOORE,  VERANUS  A.  Laboratory  directions 
for  beginners  in  bacteriology.  Cornell  Uni- 
versity (Ithaca,  N.  Y.).  2d  ed.,  revised, 
looo.  Ginn  &  Co.,  The  Athenaeum  Press, 
Boston,  Mass.  pp.  xvi,  143. 

('oo).  MIGULA,  W.     Specielle   Systematik  der  Bak- 
terien.   2  Bd.    8vo.    pp.  ix,  1,068,  with  18 
tables   from  photomicrographs  and  35  text 
figures.     Jena,  Gustav  Fischer,   1900.     (Is- 
sued two  months  ahead  of  time.) 
This  book  should  be  in  every  laboratory.    No  other 
general  work  deals  as  carefully  with  the  morphology  of 
the  bacteria.    It  is  designedly  less  complete  in  its  treat- 
ment of  cultural  characters. 


206 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('oo).  LEVY,  ERNST,  AND  KLEMPERER,  FELIX.  Ele- 
ments of  clinical  bacteriology  for  physicians 
and  students.  Second  enlarged  and  revised 
edition.  Authorized  translation  by  Augus- 
tus A.  Eshner,  Philadelphia.  W.  B.  Saun- 
ders,  1900.  pp.  441,  92  figs. 

('oi).  ABBOTT,  A.  C.  The  hygiene  of  transmissible 
diseases,  their  causation,  mode  of  dissemina- 
tion, and  methods  of  prevention.  W.  B. 
Saunders  &  Co.,  Philadelphia  and  London, 
1901.  2d  ed.  pp.  350,  with  46  ill.  and  charts. 
Popular. 

(*oi).  ALMQUIST,  E.,  OCH  TROILI-PETERSSON,  G. 
Mikroorganismerna  i  praktiska  Lifvet  Bak- 
teriologiens  utreckling  och  untidastand- 
punkt.  184  pp.  8vo.  Stockholm,  1901. 

('oi).  CHESTER,  FREDERICK  D.  A  manual  of  deter- 
minative 'bacteriology.  New  York,  The 
Macmillan  Co. ;  London,  Macmillan  &  Co., 
Ltd.,  1901.  pp.  vi,  401. 

('oi).  CONN,  HERBERT  W.  Agricultural  bacteriol- 
ogy. A  study  of  the  relation  of  bacteria  to 
agriculture,  with  special  reference  to  the 
bacteria  in  the  soil,  in  water,  in  the  dairy, 
in  miscellaneous  farm  products,  anid  in 
plants  and  domestic  animals,  pp.  vi,  412. 
Philadelphia,  P.  Blakiston's  Son  &  Co., 
1901. 

('oi).  FROST,  WILLIAM  D.  A  laboratory  guide  in 
elementary  bacteriology.  pp.  vm,  205. 
Tracy,  Gibbs  &  Co.,  Madison,  Wis.,  1901. 

('02).  International  catalogue  of  scientific  literature. 
First  annual  issue.  R.  Bacteriology.  Pub- 
lished for  the  International  Council  by  the 
Roy.  Soc.  of  London,  vol.  vm,  1902  (De- 
cember), London,  Harrison  &  Sons,  45  St. 
Martin's  Lane. 
At  least  one  additional  volume  has  beeu  published. 

('02).  BOWHILL,  THOMAS.     Manual   of  bacteriologi- 
cal technique  and  special  bacteriology.     2d 
ed.,  revised  and  enlarged.    New  York,  Will- 
iam Wood  &  Co.,  1902.    pp.  xvi,  324,  136  figs. 
('02).  EYRE,  J.  W.  H.    The  elements  of  bacteriolog- 
ical technique,   a  laboratory   guide   for  the 
medical,  dental,  and  technical  student.     170 
illustrations.     Philadelphia  and  London,  W. 
B.  Saunders  &  Co.,  1902.    pp.  371- 
('02?).  Collection   de   planches  murales   destinees   a 
I'en'seignement    de    la    bacteriologie.      Pub- 
liees  pair  1'Institut  Pasteur  de  Paris. 
Cette  collection  louche  comme  principaux  sujets  ;  char- 
bon,  rouget,  cholera  des  poules,  pneumonic,  lepre,  suppu- 
ration, peste,  gonocoque,  cholera,  fievre  typhoide,  morve, 
tuberculose.lepre,  actinomycose,  diptherie,  tetauos,  etc., 
et  les  maladies  a   protozoaires :    coccidies,  paludusme, 
maladie  de  la  mouche  tse-tse,  trypanosomes,  etc.    Condi- 
tions de  la  publication.    La  collection  cotnprend  actuel- 
lement  65  planches  du  format  Sox  62  centimetres,  tirees  en 
couleurs  sur  papier  toile  tres  fort,  munies  d'oeillets  per- 
mettant  de  les  suspendre  sur  deux  pitons  et  reuuies,  dans 
un  carton  dispose  specialement  a  cet  effet.    EHe  est  ac- 
compagnee  d'un  texte  explicatif  redige  en  trois  langnes 
(francais,  allemand,  anglais).    Prix:  250  francs  (port  en 
BUS).    Les  planches  ne  sont  pas  vendues  separement. 

(' — ).  PARK,  WILLIAM  H.  Bacteriology  in  Medicine 
and  Surgery.  Lea  Brothers  &  Co.,  New 
York.  I2mo.,  688  pages,  with  87  illustrations 
in  black  and  colors,  and  2  colored  plates. 

('02).  ROGER,  G.  H.  Les  maladies  infeotieuses.  Paris, 
Masson  et  Cie,  1902,  T.  I,  pp.  xiv,  664;  T. 
ii,  pp.  665-1,520;  av.  117  fig. 


('02).  'MATZUSCHITA,  Tfiisi.  Bacteriologische  Diag- 
npstik.  Zum  Gebrauche  in  den  bacteriolo- 
gischen  Laboratorien  und  zum  Selbstun- 
terrichte.  Fur  Aerzte,  Tierarzte  r.id  Bo- 
taniker.  Jena  (G.  Fischer),  1902,  pp.  xvn, 
692,  mit  i  Taf. 

('02).  HEWLETT,  RICHARD  T.  A  Manual  of  Bac- 
teriology, Clinical  and  Applied,  2d  Edition. 
London  (Churchill),  1902,  p.  533,  wi*h  20 
plates. 

Co2-'o4).  KOLLE,  W.,  UND  WASSERMAN,  A.,  unter 
Mitwirkung  von  54  Fachmanner.  Handbuch 
der  pathpgenen  Mikroorganismen.  Jena, 
Gustav  Fischer.  Begun  in  1902.  20  Liefer- 
ungen  to  Sept.,  1904. 

Four  large  volumes,  with  an  atlas  containing  288  photo- 
graphs.    Monographic  in  character. 

('03).  STERNBERG,  GEO.  M.     Infection  and  immunity, 

with  special  reference  to  the  prevention  of 

infectious   diseases.     G.   P.   Putnam's   Sons, 

New  York  and  London,  1903.     pp.  ix,  293. 

Popular. 

('03).  FRANKLAND,   MRS.   PERCY.     Bacteria   in   daily 
life.      Longmans,    Green    &    Co.,    London, 
New    York,    and    Bombay,    1903.     pp.    216. 
No  illustrations. 
Popular. 

('03).  WILLIAMS,  HERBERT  U.  A  manual  of  bac- 
teriology. Ninety-nine  illustrations.  3d 
ed.,  revised  and  enlarged.  Philadelphia,  P. 
Blakiston's  Son  &  Co.,  1903.  pp.  xv,  351. 

('03).  McFARLAND,  JOSEPH.  A  text-book  upon  the 
pathogenic  bacteria  for  students  of  medi- 
cine and  physicians.  With  153  illustrations, 
a  number  of  them  in  colors.  4th  ed.,  re- 
written and  enlarged.  W.  B.  Saunders  & 
Co.,  Philadelphia,  New  York,  and  London, 
1903.  pp.  629. 

('03).  MUIR,  ROBERT,  AND  RITCHIE,  JAMES.  Manual 
of  bacteriology.  Am.  ed.  (with  additions), 
revised  and  edited  from  the  third  English 
ed.  by  Norman  MacLeod  Harris.  170  illus- 
trations. Macmillan  Co.,  New  York.  Mac- 
millan &  Co.,  Ltd.,  London,  1903.  pp.  xx, 
565. 
An  excellent  book. 


IV.  Physical,   Chemical,  Zoological,   and  Botanical 
Works  of  Special  Use  to  the  Plant  Pathologist. 

('61).  GRAHAM,  THOMAS.  Liquid  diffusion  applied 
to  analysis.  Phi-los.  Trans,  of  the  Roy.  Soc., 
London,  vol.  151,  pt.  I,  1861.  pp.  183-224. 
3  figs. 

Interesting  long  paper,  in  which  the  making  of  silicate 
jelly  is  described. 

('83).  MOLISCH,  HANS.  Ueber  _  d.  _Mikrochem. 
Naohw.  v.  Nitraten  u.  Nitriten  in  d.  Pflanze 
mittelst  Diphenylamin  oder  Brucin.  Be- 
richte  der  Deutschen  Botanischen  Gesell- 
schaft,  1883.  pp.  150-155-  See  also  Bot. 
Centralb.,  Bd.  xiv,  1883,  pp.  355-356. 

('84).  GAUTIER,  ARM  AND.  Traite  de  chimie  appliquee 
a  la  physiologic  et  a  1'hygiene.  Paris,  1884. 

Not  seen . 


PHYSICAL,   CHEMICAL,   ZOOLOGICAL,   AND  BOTANICAL  WORKS. 


207 


('84).  DE  VRIES,  HUGO.  Over  eene  methode  om  in 
planten-sappen  gebonden  zuren  te  befolen. 
Maandblad  voor  Natuurwetensdiappen,  No. 
9,  1884.  Rev.  in  Botan.  Centralb.,  Bd.  xxiv, 
p.  249. 

This  author  gives  a  method  of  estimating  the  quantity 
of  acid  in  plants  when  not  free,  i.e.,  when  united  with 
bases. 

('84).  MOLL,  J.  W.  Eene  nieuwe  mikrochemische 
looizuurreactie.  Maandblad  voor  Natuur- 
wetenschappen,  1884.  Rev.  in  Bot.  Centralb. 
Bd.  xxiv,  p.  250. 

Best  method  (micro-chemical)  of  detecting  tannin  in 
cells.  He  soaks  small  pieces  of  tissue  in  saturated  solu- 
tion (7  per  cent)  acetate  of  copper  8-10  days,  and  longer 
will  not  injure.  Then  they  are  sectioned  and  treated  for 
a  few  minutes  with  a  drop  of  0.5  per  cent  sol.  iron  ace- 
tate. Too  long  an  exposure  browns  the  cell  walls.  The 
sections  are  then  washed  in  water,  transferred  to  alcohol 
and  finally  mounted  in  glycerine  or  glycerine  jelly.  If 
the  test  cannot  be  made  at  once,  the  specimens  will  not 
be  injured  by  preserving  in  alcohol,  after  the  soaking  in 
acetate  of  copper.  The  tannin-bearing  cells  become  green 
or  blue. 

('85).  HANAUSEK,  T.  P.,  AND  PAMMER,  L.  Ueber 
die  Loslichkeitsverhaltnisse  des  Kautschuks. 
Zeitschrift  des  allg.-  6'sterr.  Apotheker- 
Vereins,  1885,  No.  31,  pp.  486-488,  with  a 
table.  Rev.  in  Bot.  Centralb..  Bd.  xxv,  1886, 
P.  308. 
Note  on  best  solvents  for  rubber. 

('85)-  STRASBUKGER,  ED.  Zur  mikroskopischen 
Technik.  Bot.  Centralb.,  Bd.  xxiv,  1885, 
pp.  156-157- 

According  to  Strasburger,  Faber's  yellow  pencils  are 
the  best  for  writing  on  glass  or  porcelain.  To  find  par- 
ticular places  in  a  preparation,  be  makes  rings  on  the 
stage  of  the  microscope  to  each  side  of  the  opening,  and, 
when  the  desired  field  is  in  view,  corresponding  ones  ou 
the  slide.  The  best  method  of  making  eau  de  Javelle  is 
said  to  be  the  following  : 

(rt)  Take  20  parts  of  25  per  cent  chloride  of  lime  (Chlor- 
kalkes),  stir  up  in  100  parts  of  water,  and  let  stand  for  a 
time. 

(6)  Dissolve  15  parts  of  pure  potash  in  100  parts  of 
water. 

Add  b  to  «,  and,  after  one  to  several  days,  filter  and 
use  the  filtrate,  if  lime  still  remains  in  the  fluid,  add  a 
few  drops  of  potash  solution  and  filter  out  precipitate. 

('85).  LEHMANN,  Q.  Physikalische  Technik.  Spe- 
cielle  Anleitung  zur  Selbstanfertigung  phy- 
sikalischer  Apparate.  pp.  XH,  416,  with  882 
wood  cuts  and  17  tables.  Leipsic,  Wm. 
Engelmann,  1885. 

('85).  NOLL,   F.     Eau   de  Javelle,   ein   Aufhellungs- 
und    Losttngsmittel     fur     Plasma.      Botan. 
Centralb.,  Bd.  xxi,  1885,  pp.  377-380. 
Author  tells  how  to  make  and  how  to  use  ean  de  Javelle. 
Alcoholic  material  is  best  suited  for  treatment.    Fresh 
protoplasm  does    not    dissolve    completely,    but   leaves 
detritus  and  granules.     Strasburger,  Ibid.,  Band  xxiv, 
p.  157,  says  this  is  not  eau  de  Javelle. 

('85).  KRAUS,  C.  Ueber  amphotere  Reaction  der 
Pflanzen-Safte.  Botan.  Centralb.,  Bd.  xxiv, 
1885,  p.  287. 

A.  Meyer  summarizes  some  of  Kraus'  results  obtained 
from  the  pith  parenchyma  of  twenty  plants.  Kraus 
claims  that  neutral  litmus  would  not  answer.  He  used 
red  and  blue  litmus.  Little  pieces  were  thrown  into 
the  sap  and  left  a  long  time.  He  got  both  reactions. 

('86).  PFEFFER,  W.  Ueber  Aufnahme  von  Anilin- 
farben  in  lebende  Zellen.  Untersuchungen 
a.  d.  Bot.  Inst.  zu  Tuebingen,  Leipzig,  1886, 
Bd.  n,  Heft  2,  pp.  179-329. 

('86).  BACH  MANN,  E.  Spektroscopische  Untcr- 
suchugen  von  Pilzfarbstoffen.  Plauen,  1886. 
Not  seen. 


('86).  SHENSTONE,  W.  A.  The  methods  of  glass 
blowing  for  the  use  of  physical  and  chemical 
students,  pp.  x,  86,  with  figures.  I2mo. 
Rivingston's,  Waterloo  Place,  London,  1886. 

('89).  BEHRENS,  W.,  KOSSEL,  A.,  UND  SCHIEFFER- 
DECKER,  P.  Das  Mikroskop  und  die  Metho- 
den  der  mikroskopischen  Untersuchungen. 
pp.  vm,  315,  with  193  wood  cuts.  Braun- 
schweig, Harald  Bruhn,  1889. 

('90).  CURTMAN,  CHARLES  O.    Chemical  reagents  and 
the  spectroscope.     I2mo.     pp.  256,  with   12 
plates.    John  L.  Boland  Book  and  Stationery 
Co.,  St.  Louis,  1890. 
A  useful  little  book. 

('90).  NICKEL,  EMIL.  Die  Farbenreactionen  der 
Kohlenstoffverbindungen.  2d  ed.  pp.  VIH, 
134.  Berlin,  Verlag  von  Hermann  Peters, 
1890. 

('9i-'92).  OSTWALD,  WILHELM.    Lehrbuch  der  allge- 

meinen   Chemie.     Zweite  ganzlich   umgear- 

beitete  A-uftage.     Leipzig,  1891-1892.     2  Bd. 

The  fourth  book  of  the  second  edition  was  translated 

into  English  by  M.   M.  Pattison  Muir,  under  the  title 

"  Solutions."     London,  1891,  pp.  xiii,  316,  8  vo. 

('92).  TRIMBLE,  HENRY.  The  tannins,  pp.  168.  J. 
B.  Lippincott  Co.,  Philadelphia,  1892. 

('92).  ZIMMERMANN,  A.  Die  batanische  Mikrpteoh- 
nik.  Ein  Handbuch  der  Mikroskopischen 
Preparations-  Reaktions-  und  Tinktions- 
methoden.  Mit  63  Abbild.  im  Text.  Tubin- 
gen, 1892.  pp.  x,  278.  English  translation  by 
James  Ellis  Humphrey.  Henry  Holt  &  Co., 
New  York,  1893. 

('93).  LEA,  A.  SHERIDAN.  The  chemical  basis  of  the 
animal  body.  New  York,  Macmillan  &  Co., 
1893.  pp.  288.  Forms  Part  v  of  6th  ed.  of 
Foster's  larger  Physiology. 

('93).  BENDER,  ADOLF,  UND  ERDMANN,  HUGO.  Chem- 
ische  Praparatenkunde.  2  vols.  Bd.  i. 
Anleitung  zur  Darstellung  anorganischer 
Praparate,  von  Bender,  pp.  vm,  530,  with 
102  figures,  1893.  Bd.  n.  Anleitung  zur 
Darstellung  organischer  Praparate,  von  Erd- 
ntan.  pp.  XH,  610,  with  41  figures.  Stutt- 
gart, Ferdinand  Enke,  1894. 

('93-'95).  SCHMIDT, ERNST.  Ausfiihrliches Lehrbuch 
der  Pharmaceutischen  Chemie.  Braunsch- 
weig, Fr.  Vieweg  &  Sohn.  3d  ed.  Inorg., 
1893.  Organic,  1895.  pp.  648,  vi.  4th  ed., 
1901.  pp.  XXXIH,  1,944. 

('94).  BEHRENS,  H.  A  manual  of  microchemical 
analysis.  With  an  introductory  chapter  by 
John  W.  Judd.  With  87  illustrations,  pp. 
xxv,  246.  London  and  New  York,  Mac- 
millan &  Co.,  1894. 

('94).  LANDOLT  UND  BOERNSTEIN.  Physikalisch-chem- 
ische  Tabellen.  2d  ed.  pp.  xi,  563.  Berlin, 
Julius  Springer,  1894. 

('94).  The  National  Dispensatory.  5th  ed.  1894. 
Lea  Brothers  &  Co.,  Philadelphia  and  New 
York.  pp.  VHI,  1903. 

('94).  SCHULZ,  G.,  AND  JULIUS,  P.  Systematic  sur- 
vey of  the  organic  colouring  matters.  Trans- 
lated from  the  German  and  edited  with  ex- 
tensive additions  by  Arthur  G.  Green,  pp. 
vm,  205.  London  and  New  York,  Mac- 
millan &  Co.,  1894. 
Very  useful. 


208 


BACTERIA   IN    RELATION    TO    PLANT    DISEASES. 


('94).  LIDFORSS,  B.  Ueber  die  Wirkungssphare  der 
Glycose  u.  Gerbstoff-Reagenten.  Sep.  Abdr. 
aus  Limds  Univ.  Arsskr.  T.  xxvm,  14  pp. 
Rev.  Bot.  Centralb.,  1894,  Bd.  ux,  p.  281. 

Author  says  tannins  give  most  of  the  protein  reactions, 
and  may  thus  lead  investigators  to  many  wrong  conclu- 
sions.  The  following  test  is  given  for  grape  sugar  in 
plant  tissues :  The  parts  to  be  tested  are  soaked  for 
some  time  in  an  alcoholic  copper  solution,  and  then 
brought  to  a  boil  over  a  water  bath.  This  causes  a  pre- 
cipitate of  copper  oxide  (oxydul)  in  all  of  the  cells  which 
contained  grape  sugar.  The  alcoholic  copper  solution  is 
made  as  follows  :  Add  some  acetic  acid  and  glycerin  to 
an  alcoholic  solution  of  copper  acetate.  This  is  then 


substances  which  reduce  Feliling.  A  great  number  of 
soluble  reducing  substances  are  removed  by  the  alcohol 
from  the  plant  cells,  while  the  sugar  remains  behind. 

Barfoed's  reagent  consists  of  acetate  of  copper  dis- 
solved in  water  and  made  acid  by  acetic  acid.  It  is  useful 
for  detection  of  sugar  in  some  cases  where  Fehling's 
solution  is  worthless,  i.  e.,  in  the  presence  of  PhlorogTu- 
cin,  Aesculin,  Quercit,  which  reduce  the  latter.  The  for- 
mer, on  the  contrary,  is  reduced  by  Hydrochinon  and 
Resorcin. 

C9S).  CROSS  AND  BEVAN.  Cellulose.  An  outline  of 
the  chemistry  of  the  structural  elements  of 
plants,  with  reference  to  their  natural  his- 
tory and  industrial  uses.  pp.  vi,  320,  with 
14  plates.  London,  Longmans,  Green  &  Co., 
1895- 

('95).  MEYER,  ARTHUR.  Untersuchungen  iiber  die 
Starkekorner.  8vo.  pp.  xvi,  318,  with  9 
lithographic  plates  and  99  text  figures. 
Jena,  Gustav  Fischer,  1895. 

('95).  VON  LIPPMAN,  E.  O.  Die  Chemie  der  Zuck- 
erarten.  8vo.  Second  edition,  pp.  xxvi, 
1,174.  Braunschweig,  Fr.  Vieweg  u.  Sohn, 
1895. 

The  third  edition  appeared  in  1904  in  two  large  vol- 
umes, pp.  xxxviii,  2003.  Indispensable. 

CtS).  BEHRENS,  H.  Anleitung  zur  mikrochemischein 
Analyse  der  wichtigsten  organischen  Ver- 
bindungen.  I  Hft.  Anthracenegruppe, 
Phenole,  Chinone,  Ketone,  Aldehyde,  pp. 
viii,  64,  with  49  figures,  1895.  2  Hft.  Die 
wichtigsten  Faserstoffe.  pp.  viii,  108,  with 
18  figures  in  text  and  three  colored  plates. 
1896.  3  Hft.  Aromatische  Amine.  pp.  vn, 
135,  with  77  figures,  1896.  4  Hft.  Karba- 
mide,  aliphatische  Karbonsauren,  Aroma- 
tische Karbonsauren.  Hamburg  and  Leipsic, 
Leopold  Voss,  1897. 

Very  useful. 

('96).  BEYERINCK,  M.  W.  Kulturversuche  mit 
Amoben  auf  festem  Substrate.  Centralb.  f. 
Bakt.,  i  Abt.,  Bd.  xix,  pp.  257-267,  1896. 

('96).  COMEY,  ARTHUR  M.  A  Dictionary  of  Chem- 
ical Solubilities.  Inorganic,  pp.  xx,  515. 
Macmillan  &  Co.,  London  and  New  York, 
1896. 

('96).  Merck's  Index.  An  encyclopaedia  for  the 
physician  and  the  pharmacist;  stating  the 
names  and  synonyms ;  source  or  origin ; 
chemical  nature  and  formulas;  physical 
form,  appearance,  and  properties;  melting 
and  boiling  points;  solubilities;  gravities 
and  percentage  strengths ;  physiological  ef- 
fects ;  therapeutic  uses ;  modes  of  adminis- 
tration and  application;  regular  and  maxi- 


mum dosage;  incompatibles ;  antidotes; 
special  cautions ;  hints  on  keeping  and 
handling;  methods  of  testing;  market  val- 
ues, etc.,  of  the  chemicals  and  drugs  used  in 
medicine,  in  chemistry,  and  in  the  arts.  2d 
ed.  Merck  &  Co.,  New  York,  1896. 
Very  useful. 

('96).  LEFEVRE,  LEON.  Traite  dcs  matieres  color- 
antes  organiques  artificielles,  de  leur  prepa- 
ration industrielle  et  de  leur  applications. 
2  vols.  8vo.  pp.  xx,  1,648,  with  31  text 
figures  and  261  dyed  or  printed  samples  of 
silk,  wool,  cotton,  paper,  and  leather,  pre- 
pared under  the  direction  of  the  author 
especially  for  this  edition.  Paris,  G.  Mas- 
son,  1896. 

('97).  DAVENPORT,  C.  B.  Experimental  morphology. 
Pant  i.  Effect  of  chemical  and  physical 
agents  upon  protoplasm,  1897.  The  Mac- 
millan Co.,  New  York.  pp.  xiv,  280,  74 
figs. 

('97).  MILLER,  WALTER.  Scientific  Names  of  Latin 
and  Greek  Derivation.  Proc.  Calif.  Acad. 
Sci.  Zoology.  Third  Ser.  Vol.  i.  No.  3, 
pp.  iiS-143,  1897- 

(*97).  SCHNEIDER,  A.,  AND  ALTSCHUL,  JULIUS.  Rea- 
gents and  reactions  known  by  the  names  of 
their  authors.  Milwaukee,  Wis.,  1897. 
Pharm.  Review  Pub.  Co. 

This  book  was  first  written  by  A.  Schneider.  The  Ger- 
man edition  was  revised  and  enlarged  by  Julius  Altschul. 
It  was  translated  into  English  by  Richard  Fischer. 

('97).  BAUSCH,  EDWARD.  Manipulation  of  the  micro- 
scope. Bausch  and  Lomb  Optical  Co., 
Rochester,  N.  Y.  3d  ed.  200  pp.  1897. 

('97-'o4).  PFEFFER,  WILHELM.  Pflanzenphysiologie. 
Ein  Handbuch  der  Lehre  vom  Stoffwechsel 
und  Kraftwechsel  in  der  Pflanze.  Bd.  i. 
Stoff'wechsel.  pp.  x,  620,  mit  70  Holzschnit- 
ten.  Leipzig,  1897.  Bd.  n,  Kraftwechsel. 
i  Halite,  1901,  pp.  353,  mit  31  Holzschnitten. 
ii  Halfte,  1904,  pp.  xi,  353-986,  mit  60  Holz- 
schnitten. Verlag  von  Wilhelm  Engelmann. 
English  translation  by  Alfred  J.  Ewart, 
Oxford,  Clarendon  Press.  Vol.  i,  1900. 
Vol.  ii  (Part  i),  1903.  Vol.  in  in  prepara- 
tion. 

('98).  BEHRENS,  WILHELM.    Tabellen  zum  Gebrauch 
bei      mikroscopischen      Arbeiten.        Braun- 
schweig,  Harald   Bruhn.     pp.   VH,  237.     3d 
ed.,  revised,  1898. 
Extremely  useful. 

('98).  VON  BUNGE,  GUSTAV.  Lehrbuch  der  physi- 
ologischen  und  pathologischen  Chemie.  In 
neunundziwanzig  Vorlesungen  fur  Aerzte 
und  Studierende.  4  vermehrte  und  verbes- 
serte  Aufl.  Leipzig,  Verlag  von  F.  C.  W. 
Vogel,  1898.  pp.  iv,  510. 

English  translation,  from  4th  German  ed.  by  Starling, 
London,  1902. 

('98).    SCHROEDER    VON    DER    KoLK,    J.     L.     C.       KurZC 

Anleitung  zur  mikroscopischen  Krystal- 
bestimmung.  pp.  60,  with  text  figures. 
Wiesbaden,  C.  W.  Kreidel's  Verlag,  1898. 


PHYSICAL,   CHEMICAL,   ZOOLOGICAL,   AND  BOTANICAL  WORKS. 


209 


('98-'o3>.  ALLEN,  ALFRED  H.  Commercial  organic 
analysis.  A  treatise  on  the  modes  of  assaying 
the  various  organic  chemicals  and  products 
employed  in  the  arts,  manufactures,  medi- 
cine, etc.,  with  concise  methods  for  the  de- 
tection of  impurities,  adulterations,  etc.  8vo. 

Vol.  i.  Alcohols,  neutral  alcoholic  derivatives,  etc., 
ethers,  vegetable  acids,  starch,  sugars,  etc.  3d  ed.,  1898, 
pp.  xii,  557. 

Vol.  ir,  Part  I.  Fixed  oils  and  fats,  glycerol,  explosives, 
etc.  3d  ed.,  1899,  pp.  x,  387. 

Vol.  n,  Part  II.  Hydrocarbons,  mineral  oils,  lubricants, 
benzenes,  naphthalenes  and  derivatives,  creosotes,  phe- 
nols, etc.  3d  ed.,  1901,  pp.  viii,  330. 

Vol.  n,  Part  in.  Terpenes,  essential  oils,  resins,  cam- 
phors, etc.  3d  ed.  preparing. 

Vol.  in,  Part  I.  Tannins,  dyes,  and  coloring  matters. 
3d  ed., enlarged  and  rewritten.  Illustrated.  1900, pp.  xvi, 
589. 

Vol.  in,  Part  II.  The  amines,  hydrazines  and  deriva- 
tives, pyridine  bases,  the  antipyretics,  etc.  Vegetable 
alkaloids,  tea,  coffee,  cocoa,  etc.  8  vo.  2d  ed?,  1892, 
pp.  viii,  584. 

Vol.  ill,  Part  ill.  Vegetable  alkaloids,  non-basic  vege- 
table bitter  principles.  Animal  bases,  animal  acids, 
cyanogen  compounds,  etc.  2d  ed.,8  vo.,  1896,  pp.  xii,  508. 

Vol.  IV.  The  proteids  and  albuminous  principles.  2d 
ed.,  1898,  pp.  xi,  584. 

('99).  COHN,  ALFRED  I.  Indicators  and  test  papers, 
their  source,  preparation,  application,  and 
tests  for  sensitiveness,  etc.  New  York, 
John  Wiley  &  Sons;  London,  Chapman  & 
Hall,  Ltd.,  1899.  pp.  ix,  249. 

Very  useful. 

('99).  The  Dispensatory  of  the  United  States  of 
America.  iSth  ed.,  1899.  Philadelphia,  J. 
B.  Lippincott  Company,  pp.  XLV,  1,999. 

('99).  BEHRENS,  H.  Anlekung  zur  mikrochemi- 
schen  Analyse,  pp.  xi,  242,  with  96  figures. 
2d  ed.  Hamburg  and  Leipsic,  Voss,  1899. 

Coo).  SCHOENICHEN,  WALTER,  UND  KALBERLAH, 
ALFRED.  B.  Eyferth's  Einfachste  Lebens- 
formen  des  Tier-  und  Pflanzenreiches. 
Naturgeschichte  der  mikroskopischen  Suss- 
wasserbewohner.  3  vollstandig  neubear- 
beitete  und  vermehrte  Aufl.  Mit  iiber  700 
Abbildungen  auf  16  Tafeln  in  Lichtdruck 
nach  Zeichnungen  von  Dr.  A.  Kalberlah. 
Braunschweig,  Verlag  von  Benno  Goeritz, 
1900.  pp.  viii,  556. 

('oo).  SUTTON,  FRANCIS.     A  systematic  handbook  of 
volumetric  analysis.     8th  ed.,  enlarged  and 
improved,   1900.     pp.   xi,  640,  with   figures. 
London,  J.  and  A.  Churchill. 
This  book  should  be  in  every  laboratory. 

Coo).  VAN  RIJN,  J.  J.  L.  Die  Glykoside  chemische 
Monographic  der  Pflanzenglykoside,  nebst 
systematiscber  Darstelhmg  der  kunstlichen 
Glykoside.  Gebruder  Borntraeger,  Berlin, 
1900.  pp.  xvi,  511. 

('oo).  BEHRENS,  H.  Mikrochemische  Technik.  Ham- 
burg u.  Leipzig,  Leopold  Voss,  1900.  pp. 
vin,  68. 

('oo).  BUECHLER,  MAX.    Die  Diastasen,  1900. 
Not  seen. 

('oi).  GAGE,  S.  H.  The  Microscope.  An  introduc- 
tion to  microscopic  methods  and  to  his- 
tology. 8th  ed.,  revised,  1901.  pp.  iv,  299, 
with  over  two  hundred  figures.  Comstock 
Publishing  Co.,  Ithaca,  N.  Y. 
Deserves  a  place  in  every  laboratory. 


Cor).  COBLENTZ,  VIRGIL.  A  manual  of  volumetric 
analysis,  treating  on  the  subjects  of  indica- 
tors, test-papers,  alkalimetry,  acidimetry, 
analysis  by  oxidation  and  reduction,  iodome- 
try,  assay  processes  for  drugs  with  the 
titrimetric  estimation  of  alkaloids,  estima- 
tion of  phenol,  sugar,  tables  of  atomic  and 
molecular  weights.  P.  Blakiston's  Son  & 
Co.,  Philadelphia,  1901.  pp.  viii,  181. 

Coi).  LEE,  ARTHUR  BOLLES.  The  Microtomis.t's 
Vade-Mecum,  a  handbook  of  the  methods  of 
microscopic  anatomy.  5th  rev.  ed.  Phila- 
delphia, P.  Blakiston's  Son  &  Co.,  1901. 
PP-  xiv,  532. 

Very  useful.  French  ed  by  I,ee  and  Henneguy.  Paris, 
Octave  Doin,  1902. 

('oi).  CROSS,  C.  P.,  AND  BEVAN,  E.  J.  Researches  on 
cellulose,  1895-1900.  Longmans,  Green  & 
Co.,  London,  New  York,  and  Bombay,  1901. 
pp.  vn,  180. 

('oi).  HANAUSEK,  T.  F.  Lehrbuch  der  technischen 
Mikroskopie.  With  256  text  figures.  Stutt- 
gart, Ferdinand  Enke,  1901.  pp.  x,  456. 

('oi).  RAWSON,  CHRISTOPHER,  GARDINER,  WALTER 
M.,  AND  LAYCOCK,  W.  F.  A  dictionary  of 
dyes,  mordants,  and  other  compounds  used 
in  dyeing  and  calico  printing.  London, 
Chas.  Griffin  &  Co.,  Ltd.;  Philadelphia,  J. 
B.  Lippincott  Co.,  1901.  pp.  372. 

('02).  OM£LIANSKI,  W.  Ueberdie  Garungder  cellu- 
lose. Centralb.  f.  Bakt.,  2  Abt.,  Bd.  vin, 
Nos.  7-13,  pp.  193-201,  225-231,  257-263,  289- 
294,  321-326,  3S3-36i,  385-391,  i  text  fig.  and 
i  plate.  Jena,  1902. 

Omelianski  finds  two  morphologically  similar  bacteria 
capable  of  fermenting  pure  cellulose  (Swedish  filter  pa- 
per) in  mineral  solutions  with  chalk.  Both  grow  ansero- 
bically,  and  bear  spores  in  a  swollen  terminal  part.  One 
called  the  hydrogen  ferment  breaks  up  cellulose  with  the 
formation  of  hydrogen,  carbon  dioxide,  acetic  acid,  and 
butyric  acid.  The  other,  called  the  marsh-gas  ferment, 
breaks  up  cellulose  with  the  formation  of  marsh.gas, 
carbon  dioxide,  acetic  acid,  and  butyric  acid.  The 
cultures  were  made  by  the  selective  method,  by  which 
means  most  of  the  accompanying  forms  were  crowded 
out.  The  hydrogen  ferment  was  isolated  from  the 
methane  ferment  by  heating  the  material  used  for 
the  first  transfer  (from  the  methane-yielding  ferment) 
for  15  minutes  at  70° ,  the  subsequently  inoculated 
flasks  then  gave  only  the  hydrogen  fermentation.  The 
organism  of  the  latter  was  isolated  pure  on  potato, 
but  only  after  many  trials  and  with  feeble  growth  and 
weak  ferment  powers.  The  methane  bacterium  was  not 
obtained  pure  in  colonies.  Neither  organism  colored 
blue  with  iodine.  The  experiments  were  begun  In  1894 
and  carried  through  a  long  series  of  years,  involving  an 
enormous  amount  of  painstaking  labor. 

('02).  MANN,  GUSTAV.  Physiological  histology. 
Methods  and  theory,  pp.  xv,  488.  Oxford, 
The  Clarendon  Press,  1902. 

('02).  HOEBER,  RUDOLF.  Physikalisohe  chemie  der 
Zelle  und  der  Gewebe.  Wilhelm  Engel- 
mann,  Leipzig,  1902,  pp.  xii,  344,  with  21  fig. 

('03).  KUESTER,  ERNST.  Parhologische  Pflanzenana- 
tomie.  Verlag  von  Gustav  Fischer,  Jena, 
1903,  PP-  vii,  312,  with  121  figures. 

('03).  DAVENPORT,  CHARLES  BENEDICT.  Experimental 
Morphology.  Part  n.  Effects  of  Chemical 
and  Physical  Agents  upon  Growth.  8vo. 
The  Macmillan  Co.,  New  York. 


210 


BACTERIA   IN    RELATION    TO    PLANT    DISEASES. 


('03).  PEIRCE,  GEORGE  JAMES.  A  Text-Book  of  Plant 
Physiology.  Henry  Holt  &  Co.,  New  York, 
1903,  pp.  vi,  291. 

('03).  COHN,  ALFRED  I.  Tests  and  reagents,  chem- 
ical and  microscopical,  known  by  their  au- 
thor's names,  together  with  an  index  of  sub- 
jects, ist  ed.  383  pp.  New  York,  John 
Wiley  &  Sons;  London,  Chapman  &  Hall, 
Ltd.,  1903. 

('03).  VON  FUERTH,  OTTO.  Vergleichende  chemische 
Physiologic  der  niederen  Tiere.  Jena,  Gus- 
tav  Fischer,  1903.  pp.  XIV,  670. 

('04).  NUTTALI,,  GEORGE  H.  F.  Blood  immunity  and 
blood  relationship;  a  demonstration  of  cer- 
tion  blood-relationships  amongst  animals  by 
means  of  the  precipitin  test  for  blood,  pp. 
xn,  444.  Cambridge  (England)  University 
Press,  1904. 


V.     Books  and  Papers  of  More  or  Less  General 
Interest. 

('28).  EHRENBERG,  C.  G.  Symbols  physicae,  seu 
icones  et  descriptiones  animalium  everte- 
bratorum  seposi.tis  insectis  quae  ex  itinere 
per  Africam  Borealem  et  Asiam  Occiden- 
talem  Friderici  Guilelmi  Hemprich  et  Chris- 
taani  Godofredi  Ehrenberg  medicinae  et 
chlrurgias  doctorum  studio  novae  aut  illus- 
tratas  redierunt  percensuit  et  regis  iussu  et 
impensis  edidit  Dr.  C.  G.  Ehrenberg.  Decas 
prima.  Berolin-i  ex  Officina  Academica 
Venditur  a  mittlero  MDCCCXXVIII. 
('30).  EHRENBERG,  C.  G.  Beitrage  zur  Kenntnis  der 
Organisation  der  Infusorien,  etc.  Printed 
13  Aug.,  1830,  as  part  of  Abhandl.  d.  K. 
Akad.  d.  Wissensch.  zu  Berlin.  Physic. 
Klasse  for  the  year  1830.  As  a  whole  the 
volume  bears  date  of  1832. 

('32).  EHRENBERG,  C.  G.  Die  geographische  Ver- 
breitung  der  Infusionsthierchen  in  Nord- 
Afrika  und  West-Asien,  etc.  Abhandl.  d. 
K.  Akad.  d.  Wissensch.  zu  Berlin.  Physik. 
Klasse.  For  the  year  1829.  Printed  1832. 
('32).  EHRENBERG,  C.  G.  Ueber  die  Entwickelung 
und  Lebensdauer  der  Infusionsthiere,  etc. 
Abhandl.  d.  K.  Akad.  d.  Wissensch.  zu 
Berlin,  1831.  Printed  January,  1832. 
('38).  EHRENBERG,  C.  G.  Die  Infusionsthierchen  als 
vollkommene  Organismen.  Ein  Blick  in 
das  tiefere  organische  Leben  der  Natur, 
von  Christian  Gottfried  Ehrenberg  zu  Ber- 
lin. Nebst  einem  Atlas  von  vierundsechszig 
colorirten  Kupfertafeln  gezeichnet  vom 
Verfasser.  Leipzig,  Verlag  von  Leopold 
Voss,  1838.  pp.  548,  folio. 

('40).  HENLE,    J.      Pathplogische    Untersuchungen. 
Berlin,  August  Hirschwald.   pp.  vi,  274.    1840. 
('54).  SCHROEDER,  H.,  UND  VON  DuscH,  TH.     Ueber 
Filtration  der  Luft  in  Beziehung  auf  Faul- 
niss  und  Gahrung.    Annalen  der  Chemie  und 
Pharmacie,  Bd.  LXXXIX    (Neue  Reihe,   Bd. 
xin),  Heft  i.    pp.  232-243,  Heidelberg,  1854. 
Extremely    Interesting    historically.     These    authors 
were  the  first  to  show  that  boiled  beef  bouillon  and  sim- 
ilar easily  putrescible   fluids  could   be   preserved  indefi- 
nitely if  protected  from  the  floating  matter  of  the  air  by 
means  of  cotton  plugs.     Boiled   milk,  on   the  contrary, 
decayed  as  certainly  under  the  cotton  as  when  exposed 
directly  to  the  air,  for  reasons  unknown  to  them.    The 
exact  nature  of  this  floating  matter  of  the  air  was  not 
determined.    That  remained  for  Pasteur  to  work  out 
some  years  later  by  means  of  many  ingenious  experi- 
ments. 


('54).  COHN,  FERDINAND.  Untersuchungen  iiber  die 
Entwicklungsgeschichte  der  mikroscopischen 
Algen  und  Prize.  Der  Akademie  iibergeben 
•den  i  Mai,  1853.  Nov.  Act.  Acad.  Caes. 
Leop.-Carol.  Naturae  Curiosorum.  Vol. 
xxiv,  Pars  i,  1854.  pp.  103-256,  6  plates. 
Breslau  and  Bonn. 

('59).  SCHROEDER,  H.  Ueber  Filtration  der  Luft  in 
Beziehung  auf  Faulniss,  Gahrung  und  Krys- 
tallisation.  Annalen  der  Chemie  und  Phar- 
maoie,  Bd.  cix  (Neue  Reihe  Bd.  xxxm), 
Hit.  i,  pp.  35-52,  1859.  Leipzig  und  Heidel- 
berg. 

('60).  PASTEUR,  Louis..  Experiences  relatives  aux 
generations  dites  spontanees.  C.  R.  des  se. 
de  1'Acad.  des  Sci.,  Paris,  1860.  pp.  303-307. 

('62).  PASTEUR,  Louis.  Memoire  sur  les  corpuscles 
organises  qui  existent  dans  I'atmosphere. 
Examen  de  la  doctrine  des  generations  spon- 
tanees. Ann.  de  Chimiie  et  Physique,  1862. 
3  s;erie,  T.  LXIV,  pp.  5-110. 

('65).  DAVAINE,  C.  SUT  1'existence  et  la  recherche 
des  bacteridies  dans  la  pustule  maligne.  C. 
R.  d.  se.  et  mem.  de  la  societe  de  biol.  for 
1864,  p.  93-94  (comptes  rendus  .part). 
Paris,  1865. 

('68).  ROBERTS,  W.  CHANDLER.  On  the  occurrence 
of  organic  appearances  in  colloid  silica  ob- 
tained by  .dialysis.  Journ.  of  the  Chem.  Soc. 
of  London.  New  series,  vol.  vi,  1868,  pp. 
274-276,  2  figs,  of  fungi. 

Roberts  appears  to  have  been  the  first  to  observe  the 
growth  of  organisms  on  silicate  jelly. 

('72).  COHN,  F.  Untersuchungen  fiber  Bacterien. 
Cohn's  Beitr.  zur  Biol.  der  Pflanzen,  Bd.  i. 
2  Hft.  1872.  pp.  127-224.  i  plate. 

('73).  LISTER,  JOSEPH.  A  further  contribution  to  the 
natural  history  of  Bacteria  and  the  germ 
theory  of  fermentative  changes.  Quarterly 
Journal  of  Microscopical  Science.  1873. 
Vol.  xiii.  pp.  380-408. 

('73).  PASTEUR,  Louis.  Etudes  sur  le  vin,  ses  mala- 
dies, causes  qui  les  provoquent,  precedes 
npuveaux  pour  le  conserver  et  pour  le 
vieillir.  Deuxieme  edition  revue  et  aug- 
mentee.  Avec  32  planches  imprimees  en 
couleur  et  25  gravures  dans  le  texte.  Paris, 
1873.  F.  Savy.  pp.  iv,  344. 

('76).  PASTEUR,  Louis,  fitudes  sur  la  biere,  ses 
maladies,  causes  qui  les  provoquent,  pro- 
cede  pour  la  rendre  inalterable,  avec  une 
itheorie  nouvelle  de  la  fermentation.  12 
plates,  85  text  figures.  Paris,  1876.  Gau- 
thier-Villars.  pp.  ym,  387. 

('77).  VON  NAEGELI,  C.  Die  niederen  Pilze  in  ihren 
Beziehungen  zu  den  Infectionskrankheiten 
und  der  Gesundheitspflege.  Miinchen.  R. 
Oldenbourg,  1877.  pp.  xxxn,  285. 

('78).  LISTER,  JOSEPH.  On  the  lactic  fermentation 
and  its  bearing  on  pathology.  Transactions 
of  the  Pathological  Society  of  London. 
1878.  Vol.  xxix,  pp.  425-467- 

('79).  NENCKI,  M.,  u.  GIACOSO_,  P.  Giebt  es  Bac- 
terien oder  deren  Keime  in  den  Organen 
gesunder  lebender  Thiere?  Journ.  f.  Prakt. 
Chem..  1879.  Bd.  xx,  p.  34-44. 

('82).  MOTT,  F.  W.,  AND  HORSLEY,  V.  H.  On  the 
existence  of  bacteria,  or  their  antecedents, 
in  healthy  tissues.  Journ.  of  Physiol.,  vol. 
in,  1880-1882.  pp.  188-194. 


BOOKS  AND  PAPERS  OF  MORE  OR  LESS  GENERAL  INTEREST. 


211 


('86). 


('82).  ENGELMANN,  TH.  W.  Zur  Biologic  der  Schi- 
zomyceten.  Bot.  Zeitung,  1882,  co\  321-32* 
and  337-341- 

('84).  METSCHNIKOFP,  E.  Ueber  die  Beziehung  der 
Phagocyten  zu  Milzbrandbacillen.  Archiv 
f.  patholog.  Anat.  u.  Physiologic  u.  f.  Klin. 
Med.,  Bd.  LXXIX,  1884.  pp  502-526  2 
plates. 

('86).  BOLTON,  MEADE.  Ueber  das  Verhalten  ver- 
sclnedener  Bacterienarten  im  Trinkwasser 
Ztschr.  f.  Hyg.  Bd.  i,  1886.  pp.  76-114. 
ABBE,  E.  Ueber  Verbesserungen  des  Mikro- 
skops  mit  Hilfe  neuer  Arten  optischen 
Glases.  Sitzungsber.  der  medicin.-naturw 
Gesellschaft  zu  Jena,  1886.  Also  a  separate 
24  pp.  8vo. 

('88).  SOYKA,  J.,  UND  BANDLER,  A.  Die  Entwicke- 
lung  von  (pathogenen)  Spaltpilzen  unter 
dem  wechselseitigen  Einfluss  ihrer  Zerset- 
zungsprodukte.  Fortsohr.  d.  Med.  Bd  vi 
1888.  pp.  769-773. 

Treats  of  growth  of  bacteria  in  media  exhausted  for 
other  organisms. 

('88).  BANTI,  GUIDO.     Sulla   distruzione  dei  batterii 

neirorganismo.    Arch,  per  lo  sc.  med.,  1888. 

Vol.  xn,  pp.  191-221,  with  2  pp.  of  bibliog- 
raphy, i  plate. 
('89).  VIGNAL,  WILLIAM.    Contribution  a  1'etude  des 

Bacteriacees.       Le     mesentericus     vulgatus 

Paris,  1889. 
Coo).  KOCH,  R.     Ueber  bakteriologische  Forschung 

Berlin,    1890,    Verlag   von    August   Hirsch- 

wald.    pp.  15. 
('93).  DIXON,  H.  H.     On  the  germination  of  seeds 

in  the  absence  of  bacteria.    Soi.  Trans.  Roy. 

Dublin  Soc.,  vol.  v,  series  11,  1893-1896.    pp. 

1-4.    Review  in  Rev.  Sci.,  1894,  pp.  437-438 
(94).  KOCHS,    W.      Giebt    es    ein    Zellleben    ohne 

Mikroorgamsmen?      Biol.    Centralb.,     1894. 

No.  14,  pp.  481-491. 

Answer,  yes.  Plants  are  grown  from  sterilized  seeds 
and  kept  fourteen  months  without  contact  with  bacteria 
Plums  refused  to  decay  when  the  surface  had  been  ster- 
ilized. 

('94).  FRANKLAND.  Die  Bakteriologie  in  eiraigen 
ihrer  Beziehungen  zur  chemischen  Wissen- 
schaft.  Centralb.  f.  Bakt.,  Bd.  xv,  1894, 

pp.   IOI-II2. 

('95)-  WARD,  H.  MARSHALL.  On  the  biology  of 
Bacillus  ramosus,  a  schizomycete  of  the 
River  Thames.  Proc.  Royal  Soc.  of  Lon- 
don, vol.  LVIII,  1895.  8vo.  p.  265.  Also  a 
separate. 

('95).  PFEFFER,  W.  Ueber  Election  organischer 
Nahrstoffe.  Pringsheim's  Jahrbucher,  Bd. 
xxvin,  1895,  pp.  205-268. 

('95).  ACHARD,  CH.,  ET  PHULPIN,  E.  Contribution 
a  1'etude  de  1'envahissement  des  organes  par 
les  microbes  pendant  1'agonie  et  apres  la 
mort.  Arch,  de  med.  exper.  Tome  vn,  1895, 
pp.  25-47. 

('95).  BECO,  L.  Etude  stir  la  penetration  des  mi- 
crobes intestinaux  dans  la  circulation 
generate  pendant  la  vie.  Ann.  de  1'Inst. 
Pasteur,  T.  ix,  1895,  pp.  199-209. 

C95-'96).  NUTTALL,  GEORGE  H.  P.,  UND  THIERFELDER, 
H.  Thierisches  Leben  ohne  Bakterien  im 
Verdauungskanal.  Ztschr.  f.  physiol.  Chem. 
Bd.  xxi,  1895,  pp.  109-121,  und  Bd.  xxn, 
Hft.  i,  1896,  pp.  62-73. 


( 96).  LUNT,    JOSEPH.      On     Bacillus    mesentericus 
niger  (a  new  potato  bacillus).     Centralb.  f. 
Bakt.    2.  Abt.,  Bd.  u,  1896.    pp.  572-573. 
Motile,  liquefies  gelatin  rapidly,  produces  endospores 
copiously,  blackens  potato,  curdles  milk  with  subsequent 
solution  of  the  curd,  converts  potato  starch  into  sugar 
i.  e. ,  there  is  no  iodine  reaction  after  a  time,  but  a  copious 
reduction  of  Fehling's  solution. 

('98).  SANARELLI,    G.      Das    myxomatogene    Virus. 
Beitrag  zum  Studiutn  der  Krankheitserreger 
ausserhalb    .des     Sichtbaren.      Centralb.     f 
Bakt.,  xxni  Bd.,  1898,  pp.  865-873. 
Author  thinks  it  improbable  that  there  are  unorganized 
causes  of  infection,  and  that  therefore  certain  infectious 
diseases  must  be  due  to  organisms  too  small  to  be  visible 
to  the  human  eye,  even  when  helped  by  the  best  optical 
appliances. 

('99).  OMELIANSKY,  V.  Sur  la  culture  des  mi- 
crobes nitrificateurs  du  sol.  Arch.  d.  Sci. 
biol.  St.  Petersb.  Tome  vn.  No.  4,  1899, 
pp.  291-302. 

(  99).  STURGIS,  W.  C.    A  soil  bacillus  of  the  type  of 
de   Bary's   B.   megaterium.     Phil.   Tr.   Roy. 
Soc.   of   London.     Series    B,   vol.    191,   pp. 
147-169.    pi.  14-16,  B.  172.    London,  1899. 
Organism  described  as  Bacillus  hortulanus. 

Coo).  HOF,  A.  C.  Untersuchungen  iiber  die  Topik 
der  Alkalivertheilung  in  pflanzlichen  Gewe- 
ben.  Botanisches  Centralb.,  Bd.  LXXXIII, 
No.  9,  xxi  Jahrg.,  No.  35,  1900,  pp.  273-280. 

Coo).  SMITH,  R.  GREIG.  The  (bacterial)  clouding 
of  white  wine.  Proceedings  of  the  Linnean 
Society  of  New  South  Wales,  1900.  Part 
4,  Oct.  31.  pp.  650-658.  Also  a  separate. 

Coo).  JORDAN,  EDWIN  O.  Some  observations  upon 
the  bacterial  self-purification  of  streams. 
Jour.  Exp.  Med.,  vol.  v,  pp.  271  to  314.  Dec., 
1900.  i  plate. 

Coo).  FORD,  WM.  W.  Varieties  of  colon  bacilli 
isolated  from  man.  Montreal  Medical  Jour- 
nal, Nov.,  1900.  pp.  835-844.  Also  a  sepa- 
rate, 10  pp.  Bibliography  of  14  titles. 

Coo).  SMITH,  THEOBOLD.  Adaptation  of  pathogenic 
bacteria  to  different  species  of  animals. 
Phila.  Med.  Journ.,  May  5,  1900.  Vol.  v., 
pp.  1,018-1,022. 

Coi).  FISCHER,  ALFRED.  Ueber  Protoplasmastrulctur. 
Antwort  an  O.  Buetschli.  Archiv  f.  Ent- 
wickelungsm.  d.  Org.,  1901,  Bd.  xin,  pp.  1-33. 

('02).  JOEST,  ERNST.  Unbekannte  Infektionsstoffe. 
Centralb.  f.  Bakt.,  Abt.  i,  Bd.  xxxi,  Orig- 
inale,  1902,  pp.  361-384,  pp.  410-422.  Bibliog. 
of  58  citations. 

('03).  FORD,  W.  W.  The  classification  and  distribu- 
tion of  the  intestinal  bacteria  in  man. 
Studies  from  the  Royal  Victoria  Hospital, 
Montreal,  vol.  i,  No.  5  (Pathology  u),  May, 
1903,  95  PP- 

('03).  WINSLOW,  C.  E.  A.,  AND  NIBECKER,  C.  P.  The 
significance  of  bacteriological  methods  in 
sanitary  water  analysis.  Technology  Quar- 
terly of  Mass.  Institute  of  Technology,  vol. 
xvi,  No.  3,  Sept.,  1903,  pp.  227-239.  Also  a 
separate. 

('04).  PHILLIPS,  ORVILLE  P.  A  comparative  study  of 
the  cytology  and  movements  of  the  Cyano- 
phyceae  (Plates  xxm-xxv,  pp.  237-335. 
Bibliog.  Trans,  and  Proc.  Bot.  Soc.  Penn- 
sylvania, Vol.  i.  No.  3,  1904. 
Finds  short  cillia  on  side  walls  of  Cyanophycese. 


212 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


VI.    Important  Books  and  Papers  on  Special 
Human  and  Animal  Diseases. 

('63).  DAVAINE,  C.  Recherches  sur  les  infusoires  du 
sang  dans  la  maladie  connue  sous  le  nom  de 
sang  de  rate.  C.  R.  des  se.  de  1'Acad.  des 
sci.,  1863,  T.  LVII,  pp.  220-223,  351-353,  386- 
387. 

This  and  many  other  papers  on  anthrax  are  reprinted 
in  "1'Oeuvre  de  Davaine,"  Paris,  1889,  i  vol.  8vo. 

('64).  DAVAINE,  C.  Nouvelles  recherches  sur  la 
maladie  du  sang  de  rate  consideree  au  point 
de  vue  de  sa  nature.  C.  R.  des  se.  et  mem. 
de  la  soc.  d.  biol.  for  1863,  Paris,  1864,  pp. 
193-202. 

('70).  PASTEUR,  Louis.  Etudes  sur  la  maladie  des 
vers  a  soie,  moyen  pratique  assure  de  la 
combattre  et  d'en  prevenir  le  retour.  Tome 
I,  La  pebrine  et  la  flacherie,  pp.  xu,  322, 
illustrated,  and  Tome  n,  Notes  et  docu- 
ments, pp.  327,  Paris,  1870.  Gauthier- 
Villars. 

('76).  KOCH,  ROBERT.  Aetiologie  der  Milzbrand- 
krankheit  begrfindet  auf  die  Entwicklungs- 
geschichte  des  Bacillus  anthracis.  Cohn's 
Beitrage  z.  Biol.  d.  Pflanzen.  Bd.  n,  p.  277, 
Breslau,  1876. 

('77).  PASTEUR,  L.  fitiologie  des  maladies  charbon- 
neuses.  Archives  Veter.  T.  n,  Paris,  1877, 
pp.  668-671. 

('80).  PASTEUR,  CHAMBERLAND  ET  Roux.  Sur  1'eti- 
ologie  du  charbon.  Archives  Veter.  T.  V., 
Paris,  1880,  pp.  548-557. 

('80).  PASTEUR,  Louis.  Sur  les  maladies  virulentes, 
et  en  particulier  fur  la  maladie  appelee  vul- 
gairement  cholera  des  poules.  C.  R.  des  se. 
de  1'Acad.  des  sci.  T.  xc,  1880,  pp.  239-248. 

('81).  KOCH,  ROBERT.  Zur  Aetiologie  des  Milz- 
brandes.  Mitteil.  a.  d.  K.  Gesundheitsamte. 
Bd.  i,  1881,  pp.  49-79. 

('82).  LOEFFLER  UND  SCHUETZ.  Vorlaufige  Mittheil- 
ung  fiber  die  Arbeiten  des  K.  Gesundheit- 
samtes,  welche  zur  Entdeckung  des  Bacillus 
der  Rotzkrankheit  gefiihrt  haben.  Deutsch. 
med.  Wochenschr.  Bd.  vm,  1882,  pp.  707- 
708. 

On  the  bacillus  of  glanders.     See  Vol.  CXV,  New  Syd- 
enham  Society  for  an  English  translation. 

('83).  EBERTH,  C.  J.  Der  Typhusbacillus  und  die 
intestinale  Infection.  Vortrag  226,  Volk- 
mann's  Sammel.  Klin.  Vortrage.  Innere 
Med.  No.  77,  pp.  2,033-2,050,  with  2  figs. 
Leipzig.  Breitkopf  und  Hartel,  1883. 
Bibliography  of  13  titles. 

('83).  FEHLEISEN.  Die  Aetiologie  des  Erysipels. 
Verlag  von  Theodor  Fischer.  Berlin,  1883, 
pp.  38. 

('84).  PASTEUR,    CHAMBERLAND    ET    Roux.    Sur    la 
rage.     C.    R.    des    se.    de   1'Acad.    des    Sci. 
T.  xcvni,  Paris,  1884,  pp.  1229-1231. 
Earlier   papers   will  be    found  in  the    same  journal, 
T.  xcvni,  p.  457  ;  T.  xcv,  p.  1187,  and  T.  xcn,  p.  1259. 

('84).  LOEFFLER,  FRIEDRICH.  Untersuchungen  fiber 
die  Bedeutung  der  Mikroorganismen  fur 
die  Entstehung  der  Diphtheric  beim  Men- 
schen,  bei  der  Taube  und  beim  Kalbe. 
Mitteil.  a.  d.  K.  Gesundheitsamte.  Bd.  n, 
1884,  pp.  421-499- 

('84).  GAFFKY.  Zur  Aetiologie  des  Abdominalty- 
phus.  Mittheil.  a.  d.  K.  Gesundheitsamte, 
Bd.  n,  1884,  pp.  372-420. 


('84).  KOCH,  ROBERT.     Die  Aetiologie  der  Tuberku- 
lose.    Mitteil.  a.  d.  K.  Gesundheitsamte,  Bd. 
n,  1884,  pp.  1-88,  with  10  plates. 
See  Vol.   CXV,  New  Sydenham  Society  for  English 
translation.    This  paper  should  he  read  hy  every  student 
who  desires  to  know  how  a  good  piece  of  investigation 
is  conducted. 

('84).  NICOLAIER,  ARTHUR.  Ueber  infectiosen  Teta- 
nus. Deutsche  med.  Wochenschr.,  1884. 
Bd.  x,  pp.  842-844. 

('84).  KOCH,  ROBERT.  Vortrag  fiber  die  Cholera. 
Berliner  klin.  Wochenschr.  Nos.  31  and  32, 
pp.  477-483  and  493-503.  Deutsche  med. 
Wochenschr.  Nos.  32  and  320,  1884,  pp.  499- 
507  and  519-523.  Reprinted  also  in  Fortschr. 
der  Medicin,  Bd.  n,  Beilage,  Heft  16,  1884, 
pp.  121-134  and  Heft  17,  pp.  141-168. 

Describes  the  "  comma  bacillus,"  now  generally  recog- 
nized to  be  the  cause  of  Asiatic  cholera. 

('84).  KOCH,  R.  Ueber  die  Cholerabakterien.  Deut. 
Med.  Wochenschr.,  Bd.  x,  1884,  pp.  725-728. 

('84).  CHAUVEAU,  A.,  ET  ARLOING,  S.  Etude  ex- 
perimentale  sur  la  septicemie  gangreneuse. 
Bull,  de  1'Acad.  de  med.,  Paris,  6  mai,  1884, 
2e  serie,  T.  xm,  pp.  604-615. 

('85).  NICOLAIER,  ARTHUR.  Beitrage  zur  Aetiologie 
des  Wundstarrkrampfes.  Inaugural-Disser- 
tation, Gottingen,  1885,  W.  Fr.  Kaestner, 

pp.  31- 

('87).  NOCARD  ET  Roux.  Sur  la  culture  du  bacille 
de  la  tuberculose.  Annales  de  1'Inst.  Pas- 
teur, T.  i.,  1887,  pp.  19-29. 

('88-'9o).  Roux,  E.,  ET  YERSIN,  A.  Contribution  a 
1'otude  de  la  diphtheric.  Ann.  1'Inst.  Pas- 
teur, ler  memoire,  Tome  n,  1888,  pp.  629- 
661.  2e  memoire,  Tome  HI,  1889,  pp.  273- 
288.  3e  memoire,  Tome  iv,  1890,  pp.  385-426. 

('89).  BEHRING.  Beitrage  zur  Aetiologie  des  Milz- 
brandes.  Ztschr.  f.  Hyg.,  Bd.  vn,  1889,  pp. 

171-185. 

('89).  KITASATO,  S.  Ueber  den  Tetanus  bacillus. 
Ztschr.  f.  Hygiene,  Bd.  vii,  1889,  pp.  225- 
234,  i  plate. 

('89).  Hog  cholera.  Its  history,  nature,  and  treat- 
ment, as  determined  by  the  inquiries  and  in- 
vestigations of  the  Bureau  of  Animal  Indus- 
try, U.  S.  Dept.  Agric.  Gov't.  Pr.  Office, 
Washington,  D.  C.,  1889,  pp.  197,  pi.  16. 
Bulletin  prepared  in  great  part  by  Dr.  Theobald  Smith. 

('90).  SMITH,  THEOBALD.  On  the  influence  of  slight 
modifications  of  culture  media  on  the  growth 
of  bacteria  as  illustrated  by  the  glanders  ba- 
cillus. Journal  of  Comparative  Medicine  and 
Veterinary  Archives,  vol.  xi,  pp.  158-161. 

('91).  KITASATO,  S.  Experimented  Untersuchungen 
fiber  das  Tetanusgift.  Zeitschr.  f.  Hyg., 
Bd.  x,  1891,  pp.  267-305. 

('91).  SMITH,  THEOBALD.  Special  report  on  the 
cause  and  prevention  of  swine  plague.  Bu- 
reau of  Animal  Industry,  U.  S.  Dept.  of 
Agriculture.  Bull.  6,  1891,  pp.  166,  12  plates. 

('91).  SMITH,  THEOBALD.  Zur  Kenntniss  des  hog- 
cholerabacillus.  Centralb.  f.  Bakt,  Bd.  ix, 
pp.  253-257,  307-311,  and  339-343. 

('92).  PFEIFFER,  R.  Vorlaufige  Mittheilungen  fiber 
die  Erreger  der  Influenza.  Deut.  med. 
Wochensdir.,  Bd.  xvin,  1892,  p.  28. 

('93).  BEHRING.      Die    Geschichte    der    Diphtheric. 

Leipzig,  1893. 
Not  seen. 


SPECIAL    HUMAN    AND   ANIMAL   DISEASES. 


213 


('93).  BRIEGER,  LUDWIG,  AND  COHN,  GEORG.  Unter- 
suchungen  iiber  das  Tetanusgift.  Zeitschr. 
f.  Hyg.,  Bd.  xv,  pp.  i-io,  1893. 

As  small  a  quantity  of  the  tetanus  poison  as  0.000,23 
gra_m  would  be  a  lethal  dose  for  a  man  weighing  70  kilos. 
This  is  an  inference  based  on  experiments  with  mice. 

('93).  MOORE,  VERANUS  A.  Observations  on  the 
morphology,  biology,  and  pathogenic  prop- 
erties of  twenty-eight  streptococci  found  in 
the  investigation  of  animal  diseases.  Bu- 
reau of  Animal  Industry,  U.  S.  Dept.  of 
Agriculture,  Bull.  No.  3,  1893,  pp.  9-30. 

('93).  MOORE,  V.  A.  Pathogenic  and  toxicogenic 
bacteria  in  the  upper  air  passages  of  do- 
mesticated animals.  Bulletin  No.  3,  Bureau 
of  Animal  Industry,  U.  S.  Dept.  Agric., 
1893,  PP-  38-48. 

('93).  PFEIFFER,  R.  Die  Aebiologie  der  Influenza. 
Zeitschr.  f.  Hyg.,  Bd.  xm,  1893,  pp.  357-386, 
8  plates. 

('94).  NOVY,  F.  G.  Ein  neuer  anaerober  Bacillus  des 
malignen  Oedems.  Zeitschr.  f.  Hyg.,  Bd. 
xvn,  1894,  pp.  209-233,  2  heliotype  plates 
from  photomicrographs  by  Dr.  Pfeiffer. 

('94).  KITASATO,  S.  The  bacillus  of  bubonic  plague. 
The  Lancet,  London,  1894  (n),  pp.  428-430. 

('95).  SiEBER-ScHOUMOW,  MME.  N.  6.  Contribution 
a  1'etude  des  poissons  venimeux.  Sur  le 
Bacillus  pisoicidus  agilis,  microbe  pathogene 
pour  les  poissons.  Arch.  d.  sci.  biol.,  Tome 
in,  1895,  St.  Petersb.,  pp.  226-256,  i  colored 
plate,  7  figs. 


Author  isolated  from  dying  fish  and  from  the 
of  the  reservoir  containing  the  sick  fish,  its  inflow  an 
outflow  pipes,  an  anaerobic,  gas  forming  (COz),  motil 
short  organism  (Bacillus  pisciclduit  auilis),  pathogeni 


was  killed  by  a  heating  (5-10  min.)  in  river  water  at  68-70° 
C. ,  and  agar  or  gelatin  cultures  yielded  the  cholera  red 
reaction  with  hydrochloric  acid.  The  organism  is  also 
toxic  to  white  mice,  guinea  pigs,  rabbits,  and  dogs,  but 
not  to  pigeons. 

('98).  SHIGA,  KIYOSHI.  Ueber  den  Erreger  der 
Dysenteric  in  Japan.  Centralb.  f.  Bakt.,  i 
Abt.,  Bd.  xxin,  1898,  pp.  599-600. 

('98).  SMITH,  THEOBALD.  A  comparative  study  of 
bovine  tubercle  bacilli  and  of  human  bacilli 
from  sputum.  The  Jour,  of  Exper.  Med., 
vol.  HI,  1898,  pp.  45I-5H. 

('98).  LOEFFLER.  Bericht  der  Commission  zur  Er- 
forschung  der  Maul-  und  Klauenseuche  bei 
dem  Institut  fiir  Infektionskrankheiten  in 
Berlin.  Erstattet  an  den  Cultusrniniister 
von  dem  Vorsitzend«n  der  Commission, 
Berlin,  Aug.  12,  1898.  Deutsche  mediz. 
Wochenschr.,  1898,  No.  35,  pp.  562-564. 
Also  a  separate.  Reprinted  in  Centralb.  f. 
Bakt.,  i  Abt.,  Bd.  xxiv,  1898,  pp.  569-574. 
Organism  passes  through  a  Chamberland  filter,  and  is 
invisible. 

('98).  PETRUSCHKY,  J.  Ueber  Massenausscheidung 
von  Typhusbacillen  durch  den  Urin  von 
Typhus-Rekonvalescenten  und  die  epidem- 
iologische  Bedeutung  dieser  Thatsache. 
Centr.  f.  Bakt.  i  Abt.,  Bd.  xxm,  1898,  pp. 
577-583. 

('98).  NOCARD,  ED.,  ET  LECLAINCHE,  E.  Les  mala- 
dies microbiennes  des  animaux.  3d  ed., 
1003.  Tome  i,  pp.  n,  668 ;  Tome,  n,  pp.  645. 
Paris,  Masson  et  Cie. 


('98) .  NOCARD  ET  Roux.  Le  microbe  de  la  peri- 
pneumonie_.  Bulletin  de  la  Soc.  Central  de 
Med.  Veterinaire.  Recueil  de  Med.  Veter- 
inaire  Annexe,  Paris.  Nouvelle  sen,  T.  16. 
Mar.  24,  1898,  pp.  213-233.  See  also  the 
Veterinary  Journal,  London,  vol.  XLVII,  pp. 
147-152. 

Authors  describe  as  the  cause  of  pleuro-pneumonia  in 
cattle  an  organism  of  very  small  size,  not  visible  clearly 
even  after  staining.  The  serum  from  diseased  foci  is 
extremely  virulent,  but  it  is  impossible  to  cultivate  any- 
thing from  this  serum  by  any  of  the  ordinary  methods. 
Many  bacteriologists  have  tried  and  failed,  including 
Nocard  and  Roux.  These  authors  finally  succeeded  in 
cultivating  it  in  bouillon  in  collodion  sacks.  The  bouil- 
lon was  inoculated  with  a  little  of  the  virulent  serum 
and  the  sacks  were  then  placed  in  the  peritoneum  of 
rabbits,  where  they  were  allowed  to  remain  some  weeks 
subject  to  osmosis.  The  organism  clouds  the  bouillon 
slightly,  and  is  visible  under  high  magnifications,  in 
bright  light,  as  innumerable,  minute,  bright,  mobile 
points.  This  bouillon  is  capable  of  reproducing  the  dis- 
ease, but  is  free  from  bacteria  cultivable  on  ordinary 
media.  Check  sacks  incubated  in  the  peritoneum  gave 
no  such  result,  neither  did  sacks  inoculated  with  heated 
virus.  Collodion  sack  cultures  incubated  in  the  perito- 
neal cavity  of  animals  were  first  used  (?)  by  Metchnikoff 
Roux  and  Salimbeni  in  their  study  of  the  toxin  and  anti- 
toxin of  cholera. 

The  authors  finally  succeeded  in  cultivating  this  or- 
ganism outside  of  the  animal  body,  by  using  a  special 
bouillon  and  a  special  agar  (see  Les  maladies  micro- 
biennes des  animaux,  1903,  T.  I.,  p.  450).  "  Virulent  albu- 
minous liquids,  pulmonary  serum  not  diluted,  or  the 
Martin  serum  bouillon  filtered  through  Chamberland  or 
Berkefeld  bougies,  gives  a  sterile  filtrate.  On  the  con- 
trary, after  dilution  of  the  same  liquids  in  a  non-albu- 
minous medium,  the  microbe  passes  through  the  Berke- 
feld and  the  Chamberland  bougie.  Under  these  condi- 
tions the  filtration  enables  one  to  obtain  without  diffi- 
culty a  characteristic  pure  culture,  even  from  impure 
products." 

('98).  NOCARD  ET  Roux.  Le  microbe  de  la  peri- 
pneumoniie.  Ann.  de  1'Inst.  Pasteur,  1898, 
T.  XH,  pp.  240-262. 

('99).  GELPKE,  THEODOR.  Bacterium  septatum  und 
dessen  Beziehungen  zur  Gruppe  der  Diph- 
therienbacterien(B.  diphtheriae  [KIebs-L6f- 
fler],  B.  pseudodiphtheriticum  [Loftier]  und 
B.  xerosis).  Arb.  a.  d.  Bact.  Institut  der 
techn.  Hochschule  zu  Karlsruhe,  n  Bd.,  2 
Hefte,  1899,  pp.  71-148.  5  plates  (40  pho- 
tomicrographs) and  4  charts.  Bibliog.  of 
45  titles. 

('oo).  WELCH,  WILLIAM  H.  Morbid  conditions 
caused  by  the  Bacillus  aerogenes  capsulatus. 
Phila.  Med.  Journ.,  vol.  vi,  1900,  pp.  202-216. 

('oo).  FLEXNER,  SIMON.  On  the  etiology  of  tropical 
dysentery.  Phila.  Med.  Journ.,  vol.  vi,  1900, 
pp.  414-424- 

The  author  calls  special  attention  to  Shiga's  results. 
Flexner  studied  this  disease  in  the  Philippines.  He  says  - 
"  That  the  bacillus  is  identical  with  the  organism  ob- 
tained by  Shiga  in  the  epidemic  of  dysentery  which  pre- 
vailed in  Japan,  there  can  be  no  reasonable  doubt.  In 
morphological,  cultural,  and  pathogenic  characteristics 
the  two  organisms  are  indistinguishable." 

('oo).  GORHAM,  F.  P.  The  gas-bubble  disease  of 
fish  and  its  cause.  U.  S.  Fish  Commission, 
Bull,  for  1899,  pp.  33-37,  1900,  Washington. 

('oo).  SMITH,  R.  GREIG.  A  new  bacillus  pathogenic 
to  fish.  Proceedings,  Linnean  Soc.,  New 
South  Wales  for  1900.  Sydney,  1901,  vol. 
xxv,  pp.  122-130.  Two  heliotype  plates. 

This  Is  named  Bacillus  piscidus  bipolaris,  in  allusion 
to  the  bipolar  germination  of  its  spores.  The  organism 
is  motile,  and  liquefies  gelatin. 


214 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Coo).  SMITH,  R.  GREIG.  A  fish  disease  from 
George's  River.  Proc.  Linnean  Soc.,  New 
South  Wales  for  1900.  Sydney,  1901,  vol. 
xxv,  pp.  605-609. 

('99-'oo).  MOSCHCOWITZ,  ALEXIS  V.  Tetanus.  A 
study  of  the  nature,  excitant,  lesions,  symp- 
tomatology, and  treatment  of  the  disease, 
with  a  critical  summary  of  the  results  of 
serum  therapy.  Studies  from  the  Dept.  of 
Pathology  of  the  College  of  Physicians  and 
Surgeons,  Columbia  University,  New  York. 
Vol.  vii,  88  pp.  A  bibliography  of  337  titles. 
1899-1900. 

('01).  SCHULTZ,  N.   K.     De  la  vitalite   du   microbe 
de    la   peste    bubonique    dans    les   cultures. 
Arch,  des  Sci.  Biol.  publiees  par  1'Inst.  Im- 
perial  de   Med.   Exper.   a   St.    Petersbourg. 
T.  vin,  No.  4,  1901,  pp.  373-389.    i  plate. 
Organism  not  sporiferous.    Author  made  various  tests 
from  old  cultures  kept  in  sealed  tubes  of  bouillon,  and 
concludes  that  under  favorable  conditions  the  pest  bacil- 
lus may  preserve  its  vitality  and  virulence  for  four  years. 
In  its  most  modified  form,  he  says,  it  exists  in  the  state 
of  very  small  round  grains. 

('01).  TARTAKOWSKY,  G.,  ET  DCHOUNKOWSKY.  Du 
microbe  de  la  peripneumonie  des  boeufs. 
Arch,  des  sci.  biol.  publiees  par  1'inst.  imper. 
de  med.  exper.  a  St.  Petersbourg.  T.  vin, 

1901,  pp.  441-460,  6  figs. 

Confirms  the  work  of  Nocard  and  Roux. 

('02) .  MARSH,  M.  C.  Bacterium  trutta?,  a  new  species 
of  bacterium  pathogenic  to  trout.  Science, 
n.  s.,  vol.  xvi,  No.  409,  pp.  706-707,  Oct.  31, 

1902.  Also  a  separate. 

This    organism   browns   nutrient    agar.    Its   thermal 
death  point  is  said  to  be  42°  C. 

('02).  SHIGA,  K.  Weitere  Studien  iiber  den  Dysen- 
teriebacillus.  Zeitschr.  f.  Hyg.  Bd.  XLI, 
1902,  pp.  3SS-368. 

('02).  MOORE,  V.  A.  The  pathology  and  differential 
diagnosis  of  infectious  diseases  of  animals. 
Ithaca,  N.  Y.,  1902.  pp.  xrv,  380.  73  figs. 
8  plates. 

('02).  KOCH,  ROBERT.  An  address  on  the  transfer- 
ence of  bovine  tuberculosis  to  man.  Brit. 
'Med.  Jour.,  London,  1902,  vol.  2,  pp.  1,885- 
1,889. 

('03).  MARSH,  M.  C.  A  more  complete  description 
of  Bacterium  truttae.  Bull.  U.  S.  Fish  Com- 
mission for  1902.  Washington,  Govt.  Print- 
ing Office,  1903,  pp.  4H-4IS,  with  two  plates. 
Also  a  separate. 

Grows  well  in  media  which  is  neutral  or  +  5,  but  there 
is  little  or  no  growth  when  the  acidity  is  -f- 15.  Growth 
is  also  inhibited  by  an  alkalinity  of  —  5.  Gelatin  and 
blood  serum  are  liquefied.  There  is  no  growth  on  potato 
unless  it  is  first  neutralized  ;  then  there  is  a  scanty  white 
growth.  Milk  is  not  coagulated,  but  it  becomes  fairly 
transparent  after  two  weeks.  It  does  not  ferment  glu- 
cose, lactose  or  saccharose.  It  reduces  nitrates  to  nitrites 
and  to  ammonia.  It  is  not  clearly  mobile  It  does  not  pro- 
duce indol  or  phenol.  The  optimum  temperature  is  at 
or  near  20°  C .  It  is  actively  pathogenic  to  trout,  especially 
brook  trout,  in  which  the  disease  was  first  observed. 

('03).  MONFALLET,  D.  Bibliographic  abregee  des 
infections.  Paris  et  Santiago  (Chili),  Ch. 
Goffi,  editeur,  1903,  pp.  1-65. 

About  1900  classified  references  to  papers  on  human 
and  animal  diseases  of  a  communicable  nature. 

('04).  REMUNGER,  P.  Les  travaux  recent  sur  la  rage. 
Bull,  de  1'Inst.  Pasteur,  1904,  T.  n,  pp.  753- 
764. 


"  We  have  shown  that  if  one  filters  an  emulsion  of 
rabies  virus  through  a  Berkefeld  V  bougie  and  then  cen- 
trifuges the  filtrate,  the  upper  layers  are  deprived  of  vir- 
iileuce,  but  the  virulence  is  kept  in  the  lower  layers. 
Barratt  lias  obtained  the  same  results  with  an  unfiltered 
emulsion  of  a  rabid  brain." 

('04).  FLEXNER,  SIMON,  HOLT,  L.  EMMETT,  and  as- 
sistants. Bacteriological  and  Clinical  Studies 
of  the  Diarrheal  Diseases  of  Infancy,  with 
Reference  to  the  Bacillus  Dysenteriaa 
(Shiga).  Studies  from  the  Rockefeller  In- 
stitute for  Medical  Research,  New  York. 
Vol.  n,  1904,  pp.  7-202. 


VII.     Predisposition,  Conditions  Favoring  Infection 
or  Immunity. 

('80).  CHAUVEAU.    See  xxn. 

('81).  LOEFFLER,  FRIEDRICH.  Zur  Immunitatsfrage. 
Mittih.  a.  d.  K.  Gesundheitsamte.  Bd.  i, 
iSSi,  pp.  134-187.  Also  a  separate,  54  pp. 

('87).  METCHNIKOFF.    See  xxi. 

('88).  NUTTALL.     See  xxxvm. 

('88).  NUTTALL.    See  xxi. 

('88).  FLUEGGE,  C.  Studien  iiber  die  Abschwachung 
virulenter  Bakterien  und  die  erworbenc- 
Immunitat.  Zeitsch.  f.  Hygiene.  1888.  Bd. 
IV,  pp.  208-230. 

('89).  BUCHNER.    See  xxi. 

('89).  NISSEN.    See  xxi. 

('90).  LEHMANN.    See  xxi 

('91).  OGATA.    See  xxi. 

('91).  BUCHNER,  HANS.  Ueber  Immunitat,  deren 
natiirliches  Vorkommen  und  kunstliche 
Erzeugung.  Munch.  Med.  Wochenschr. 
1891,  38  Jahrg.,  pp.  5SI-SS4,  574-579- 

('94).  EHRLICH  AND  WASSERMANN.    See  xxi. 

('01).  HUEPPE,  FERDINAND.  Perlsucht  und  Tuber- 
culose.  Berliner  klin.  Wochenschrift,  1901, 
No.  34,  pp.  876-878.  Also  a  separate,  pp.  10. 

(*oi).  ZABOLOTNY,  D.  Recherches  stir  la  peste.  2 
mem.  Experiences  d'inoculation,  d'im- 
munisation  et  de  traitemeiit  des  animaux. 
Arch,  des  Sci.  Biol.  publiees  par  1'Inst.  Im- 
perial de  Med.  Exper.  a  St.  Petersbourg. 
T.  vni,  No.  4,  looi,  pp.  390-427.  2  plates. 

('01).  METCHNIKOFF,  ELIE.  L'immunite  dans  les 
maladies  infectieuses.  Paris  (Masson), 
1901,  pp.  ix,  600,  av.  fig.  German  trans,  by 
Julius  Meyer.  Jena  (Fischer),  1902,  pp.  xi, 
456. 
Not  seen. 

('02).  AMMON,  OTTO.  Theoretische  Betrachtungen 
iiber  Ansteckung  und  Disposition.  Arch.  f. 
Hyg.  Bd.  XLII,  1002,  pp.  289-305. 

('o2-'o4).  KOLLE  AND  WASSERMAN.    See  in. 


VIII.    Symbiosis,  Antagonism. 

('81).  KERN,  EDWARD.     Ueber   ein   neues   Milchfcr- 
ment  aus  dem  Kaukasus.     Bull,  de  la  Soc. 
Imp.  d.  Nat.  de  Moscou,  Tome  LVI,  pp.  141- 
177,  2d  part  (No.  3),  2  plates,  1881. 
Kephir.    Dispora  caucasica,  nov.  gen.  and  n.  sp.  Plates 
good. 

(.'84).  KRANNHALS,  H.  Ueber  das  Kumys-ahnliche 
getrank  "Kephir"  und  ueber  den  "Kephir  - 
pilz.  Deutsch.  Archiv.  f.  klin.  med.  Bd. 
xxxv,  pp.  18-37,  i  plate.  Bibliography  of  18 
titles. 


SYMBIOSIS;  CARRIERS  OF  INFECTION;  MORPHOLOGY. 


215 


('84).  STRUVE,   UEINRICH.     Ueber   Kephir.     Ber.   d. 

deutsch.    chem.    Gesellsch.     Bd.    xvn,    1884, 

PP-  3M-3i6  and  1,364-1,368. 
('87).  METCHNIKOFK.    See  xxi. 
('87.  GARRE,    C.     Ueber    Antagonisten     unter    den 

Bacterien.     Correspondenzbl.    f.    Schweizer. 

/.oo^    ,TAerzte-  J?ihrS-  xvn'  l887,  PP.  385-392. 

(  88).  NUTTALL.    See  xxi. 

('88).  DE  FREUDENREICH,  E.  De  1'anitagonisme  des 
bacteries  et  de  1'immunite  qu'il  confere  aux 
milieux  de  culture.  Ann.  de  1'Inst.  Pasteur, 
T.  n,  1888,  pp.  200-206. 

('88).  DUBOIS.     See  xxvn. 

('88).  HKRICOURT.    Des     associations     microbiennes. 

Rev.  de  med.,  T.  — ,  1888,  pp.  — . 
Not  seen. 

('88).  SIROTININ.  Ueber  die  entwicklungshemmen- 
den  Stoffwechselproduote  der  Bacterien  und 
die  sog.  Retentionshypothese.  Zeitsch.  f. 
Hyg.  Bd.  iv,  1888,  pp.  262-290. 

('89).  BEYERINCK,  M.  W.  Sur  le  kefir.  Arch.  neer. 
des  sci.  ex.  et  nat,  T.  xxm,  1889,  pp.  428- 
444-  i  fig. 

('89).  ROGERS,  G.  H.  Quelques  effets  des  associa- 
tions microbiennes.  C.  R.  hebd.  d.  se.  et 
mem.  de  la  soc.  de.  biol.,  19  Janvier,  1889, 
Paris,  se.  9,  T.  i,  pp.  35-38. 

Two  bacteria,  inoffensive  to  a  given  animal,  may  be- 
come pathogenic  when  inoculated  together. 


('90). 
('91). 
('93). 

('94). 

('96?) 
('97). 
Coo). 


BLAGOVESTCHENSKY,  N.  Sur  1'antagonisme 
entre  les  bacilles  du  charbon  et  ceux  du 
pus  bleu.  Ann.  de  1'Institut  Pasteur,  T.  iv, 
1890,  pp.  689-715. 

Mix,  CHARLES  L.  On  a  kephir-like  yeast 
found  in  the  United  States.  Proc.  Amer. 
Acad.  of  Arts  and  Sciences,  1891,  n.  s.  vol. 
xvm,  pp.  102-114. 

WARD,  H.  MARSHALL.  The  ginger-beer  plant, 
and  the  organisms  composing  it.  A  contri- 
bution to  the  study  of  fermentation-yeasts 
and  bacteria.  Phil.  Trans.  Roy.  Soc.  (B) 
for  1892.  London,  1893,  vol.  183,  pp.  125  to 
197,  PI.  6. 

GALTIER,  V.  Nouvelle  recherches  sur  1'influ- 
ences  des  associations  bacteriennes.  Exalta- 
tion de  la  virulence  de  certain  microbes. 
Accroissement  de  la  receptivite.  C.  R.  des 
se.  de  1'Acad.  d.  sci.,  T.  cxvin,  1894,  pp. 
1,001-1,004. 

.  VON  FREUDENREICH,  EDUARD.  Bakteriologische 
Untersuchungen  fiber  den  Kefir.  Landw. 
Jahrbuch  d.  Schweiz,  1896.  Bd.  x,  pp.  1-20. 
2  text  figs,  and  i  heliotype  plate. 

VON  FREUDENREICH,  ED.  Bakteriologische  Un- 
tersuchungen  fiber  den  Kefir.  Centralb.  f. 
Bakt.,  2  Abt.  Bd.  in,  1897,  pp.  47-54,  2  figs. ; 
87-95,  I35-I4L 

KRAUSE.     See  xv. 


IX.    Carriers  of  Infection. 

('88).  ALESSI,  GUISEPPE.  Sulla  trasmissibilita  dei 
germi  infettivi  mediante  le  deiezioni  delle 
mosche.  Arch,  per  lo  sci.  med.  Vol.  xn, 
Torino,  1888,  pp.  279-292. 

('91).  WAITE,  M.  B.  Results  from  recent  investiga- 
tions in  pear  Wight.  Proc.  Am.  Asso.  Adv. 
Sci.,  4Oth  meeting,  Salem,  1892. 

Mr.  Waite  obtained  his  first  results  in  iSyi,  and  called 
attention  to  them  at  the  Washington  meeting  of  the  Am. 
Asso.  Adv.  Sci.  that  summer. 


( 94).  Yersen,    La   peste   bubonique    a    Hong-Kong. 
Ann.  de  1'Inst.     Pasteur.     T.  vni,  1894,  pp. 
662-667. 
The  pest  is  carried  by  rats. 

('95).  SMITH,  ERWIN  F.    Bacillus  tracheiphilus,  etc. 
Centralb.  f.  Bakt.,etc.     2  Abt.     i  Bd.,  1895, 
P-  365. 
The  disease  is  spread  by  beetles. 

('96).  SMITH,  ERWIN  F.    A  bacterial  disease  of  the 
tomato,  egg  plant,  and  Irish  potato.    Wash- 
ington, 1896,  p.  22,  and  PI.  n,  fig.  3. 
Disease  communicated  by  beetles. 

('97).  MARPMANN.     See  XLIX. 

('97).  SMITH,  ERWIN  F.  Pseudomonas  campestris 
(Parnmel).  The  cause  of  a  brown  rot  in 
cruciferous  plants.  Centralb.  f.  Bakt.  2  Abt., 
Bd.  HI,  pp.  409-410. 

Disease  communicated  by  slugs  and  by  larva:   of  the 
cabbage  butterfly. 

('98).  SIMOND,  P.  L.  La  propagation  de  la  peste. 
Ann.  de  1'Inst.  Pasteur,  T.  xn,  1898,  pp. 
625-687.  5  figs. 

('98).  NUTTALL,  GEORGE  H.  F.  Zur  Aufklarung  der 
Rolle,  welche  stechende  Insekten  bei  der 
Verbreitung  von  Infektionskrankheiten 
spielen.  Centralb.  f.  Bakt.,  Bd.  xxm,  i  Abt., 
1898,  pp.  625-635. 

('99).  NUTTALL,  G.  H.  F.  On  the  role  of  insects, 
arachnids,  and  myriapods  as  carriers  in  the 
spread  of  bacterial  and  parasitic  diseases  of 
man  and  animals.  Johns  Hopkins  Hosp. 
Repts.,  vol  vni,  No.  1-2,  pp.  1-154.  3  plates. 
Bibliography  of  366  titles. 
Noticed  in  Nature,  Dec.  14, 1899. 

Coo).  GALLI-VALERIO,  BRUNO.  Les  puces  des  rats  et 
des  souris  jouenit-elles  un  rok  important 
dans  la  transmission  de  la  peste  bubonique 
a  1'homme?  Centralb.  f.  Bakt.,  i  Abt,  Bd. 
xxvn,  1900,  pp.  1-4.  3  figs. 

('02).  GALLI-VALERIO,  BRUNO..    The  part  played  by 
the  fleas  of  rats  and  mice  in  the  transmission 
of  bubonic  plague.    Jour.  Trop.  Med.,  Lon- 
don, vol.  v,  1902,  pp.  33-36. 
.  BRENNER,  W.     Die   Schwarzfaule   des   Kohls. 

Centralb.  f.  Bakt.    2  Abt.    Bd.  xn,  p.  729. 
Disease  said  to  be  communicated  by  aphides. 


X.    General  Morphology  of  the  Bacteria.    Cytology. 

('74).  BILLROTH,  THEODOR.  Untersuchungen  fiber  die 
Vegetationsformen  von  Cocco-bacteria  sep- 
tica  und  den  Antheil  welchen  'sie  an  der 
Entstehung  und  Verbreitung  der  accidentel- 
len  Wundkrankheken  haben.  Berlin,  1874. 
Verlag  von  Georg  Reimer.  Quarto,  pp.  xrv, 
244.  5  plates. 

('77).  BILLROTH,  TH.,  AND  EHRLICH,  F.  Unter- 
suchungen fiber  Coccobacteria  septica. 
Archiv.  f.  klinische  chir.  Bd.  xx,  pp.  403- 
433,  I  pi.,  Berlin,  1877. 

('78).  HALLIER,  ERNST.  Die  Plastiden  der  niederen 
Pflanzen,  ihre  selbststandige  Entwickelung, 
ihr  Eindringen  in  die  Gewebe,  und  ihre 
verherende  Wirkung.  Leipzig.,  1878,  pp. 
92,  4  plates. 

('82).  ZOPF,  W.  Zur  Morphologic  der  Spaltpflanzen. 
Spaltpilze  und  Spaltalgen,  Leipsic,  1882,  pp. 
vi,  74.  Verlag  von  Veit  and  Comp.  7  plates. 


2l6 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('86).  HUEPPE,  FERDINAND.     Die  Formen   der   Bak- 

terien   und   ihre   Beziehungen   zu   den   Gat- 

tungen  und  Anten.     1886.     pp.  vm,  152,  24 

figs.     Wiesbaden.     C.  W.  Kreidel's  Verlag. 

('88).  POMMER,  GUSTAV.    Ein  Beitrag  zur  Kenntniss 

der   fa-denbildenden   Bakterien.     Mitth.  hot. 

Itist.    Graz,  Jena,  1888,  pp.  93-112.     I  plate. 

Bacillus  brassicse,  a  sporiferous  form,  isolated  from  an 

imperfectly  sterilized  decoction  of  cabbage  leaf. 

('89).  METCHNIKOFF,  EL.    Contributions  a  1'etude  du 

pleomorphisme  des  baoteries.    Ann.  de  1'Inst. 

Pasteur,  T.  in,  1889,  pp.  61-68.    i  plate. 
(•80).  ERNST,  PAUL.    Ueber  Kern-und  Sporenbildung 

in  Bakterien.  Zeitschr.  f.  Hyg.    Bd.  v,  1889, 

pp.  428-486.    2  plates. 
('89).  BABES,  VICTOR.  Ueber  isolirt  farbbare  Antheile 

von   Bakterien.     Zeitsohr.    f.   Hyg.     Bd.   v, 

1889,  pp.  173-190.    i  plate. 
('89)    WINOGRADSKY,    S.      Pleomorphisme    des   bac- 

teries.    Ann.  de  1'Inst.  Pasteur,  T.  in,  1889, 

pp.  249-264. 
('89).  METCHNIKOFF,  EL.    Note  sur  le  pleomorphisme 

des  bacteries.     Ann.   de  1'Inst.   Pasteur,  T. 

in,  1889,  pp.  265-267. 
('90).  ALMQUIST,   E.     Untersuchungen   fiber    einige 

Bakteriengattungen  mit  Mycelien.     Zeitsch. 

f.  Hyg.    Bd.  vin,  1890,  Heft  2,  pp.  189-197. 
('90).  BUTSCHLI,  O.    Ueber  den  Bau  der  Bakterien 

und   Verwandter   Organismen.     Leipzig,   C. 

F.  Winter,  1890,  pp.  37.    i  pi. 

Maintains  the  existence  in  the  bacteria  of  a  body  cor- 
responding to  a  cell  nucleus. 

('91).  FAMINTZIN,    A.      Eine    neue    Bakterienform, 
Newskia  ramosa.    Bulletin  de  1'Acad.  de  St. 
Petersbourg.     Nouvelle  serie   (n),  1891,  T. 
xxxiv,  p.  481. 
Not  seen . 

('91).  DANGEARD.    See  xxin. 

('91).  PROTOPOPOFF.  Sur  la  question  de  la  structure 
des  baoteries.  Ann.  de  1'Inst,  Pasteur,  1891, 
Tome  v,  p.  332-336. 

fgi)  ZETTNOW,  E.  Ueber  den  Bau  der  Bakterien. 
Centralb.  f.  Bakt.  x  Bd.,  1891,  pp.  689-694. 
i  Tafel. 

('92)  ZUKAL,  HUGO.  Ueber  den  Zellinhalt  der  Schi- 
zophyten.  Ber.  d.  deutsch.  bot.  Gesellsch. 
x,  1892,  pp.  5I-S5- 

('92) .  SJOEBRING,  NILS.    Ueber  Kerne  und  Theilung- 
en  bei   den   Bakterien.     Centralb.    f.   Bakt., 
xi  Bd.,  1892,  pp.  65-68,  with  i  colored  Tafel. 
"In  the  bacterial  body  there  may  be  demonstrated  con- 
sequently   as  in  other  cells,  two  components— nucleus 
and  cell-body,  which,  however,  can  not  always  be  d 
tinguished." 

('92).  THAXTER,  ROLAND.    On  the  Myxobacteriaceae, 

a  new  order  of   Schizomycetes.     Botanical 

Gazette,  vol.  xvn,  No.  12,  pp.  389-406.     Pis. 

xxn-xxv,  1892. 
('92).  SAUVAGEAU,   C.,  ET  RADAIS.     Sur   les   genres 

Cladothrix,    Streptothrix,    Actinomyces,    et 

description   de   deux   Streptothrix  nouveau. 

Ann.de  1'Inst.  Pasteur.    T.  vi,  1892,  pp.  242- 

273.    i  plate. 
('92) .  FoERSTER,  F.    Ueber  eine  merkwurdige  Erschei- 

nung   bei    Chromatium    Okenii    Ehrbg.    sp. 

Centralb.  f.  Bakt.,  xi  Bd.,  1892,  pp.  257-264, 

mit  I  taf.  (colored). 

Author's  figures  show  curious  bands  or  bridges  con- 
necting the  bacterial  cells  sidewise  as  well  as  end  to  end. 


('92).  TRAMBUSTI,  A.,  AND  GALEOTTI,  G.  Neuer 
Beitrag  zitrn  Studium  der  inneren  Struktur 
der  Bakterien.  Centralb.  f.  Bakt.,  xi  Bd., 
1892,  pp.  717-722,  mit  i  Taf.  (colored). 

('92).  KLEIN,  E.  Zur  Geschicbte  des  Pleomorphis- 
mus  des  Tuberculoseerregers.  Centralb.  f. 
Bakt.,  xn  Bd.,  1892,  pp.  905-906. 

('92).  BuETSCHLi,  O.  Untersuchungen  fiber  mikro- 
skopische  Schaume  und  das  Protoplasma. 
4to,  mit  6  lithogr.  Taf.  u.  23  Fig.  im  Text, 
so  wie  einem  Atlas  von  19  Mikrophoto- 
graphieen.  Leipzig,  1892.  Rev.  in  Centralb. 
f.  Bakt.,  Bd.  xm,  1893,  pp.  436-438. 

Again  maintains  the  existence  in  bacteria  of  a  Central- 
koerper  corresponding  to  a  nucleus,  and  points  out  that 
Alfred  Fischer's  criticism  of  his  former  statements  has 
no  substantial  basis.  See  especially  pp.  75  to  79. 

('94).  BEYERINCK,  M.  W.  Ueber  die  Natur  der 
Faden  der  Papilionaceenknollchen.  Centralb. 
f.  Bakt.,  Bd.  xv,  1894,  pp.  728-732. 

('94).  ILKEWICZ,  W.  Ueber  die  Kerne  der  Milz- 
brandsporen.  Centralb.  f.  Bakt.,  Bd.  xv, 

1894,  pp.  261-267,  mit  i  Figur. 

('95).  WARD,  H.  MARSHALL.  The  formation  of  bac- 
terial colonies.  Annals  of  Botany,  vol.  ix, 

1895,  pp.  653-657. 

('95).  WARD,  H.  MARSHALL.  A  false  bacterium.  An- 
nals of  Botany,  vol.  ix,  1895,  pp.  657-658. 

('9S)-  WAGER,  HAROLD.  Preliminary  note  upon  the 
structure  of  bacterial  cells.  Annals  of 
Botany,  vol.  ix,  1895,  pp.  659-661. 

('95).  COPPEN-JONES,  A.  Ueber  die  Morphologic  und 
systematische  Stellung  des  Tuberkelpilzes 
und  liber  die  Kolbenbildung  bei  Aktinomy- 
kose  und  Tuberkulose.  Centralb.  f.  Bakt., 
xvn  Bd.,  1895,  pp.  1-16  and  70-76,  with  i 
plate. 

('95).  BRUNS,  HAYO.  Ein  Beitrag  zur  Pleomorphie 
der  Tuberkelbacillen.  Centralb.  f.  Bakt., 
xvn  Bd.,  1895,  pp.  817-826,  with  8  figs. 

('95).  LUBINSKI,  Ws.  Zur  Kultivierungsmethode, 
Biologic  und  Morphologic  der  Tuberkel- 
bacillen. Centralb.  f.  Bakt.,  xvm  Bd.,  1895, 
pp.  125-128. 

('95).  BABES,  V.     Beobachtungen  fiber  die  metachro- 
matischen  Korperchen,  Sporenbildung,  Ver- 
zweigung.     Kolben-und  Kapselbildung  path- 
ogener   Bakterien.     Zeitschr.    f.    Hyg.     Bd. 
xx,  1895,  pp.  412-437,  2  plates. 
Fig.  18  e  of  Table  XI  is  particularly  instructive.    This 
shows  a  true  branching  in  the  anthrax  organism,  but  it  is 
confined  to  the  capsule. 

COPPEN-JONES,    A.      Ueber    die    Nomenclatur 

des    sog.     "  Tuberkelbacillus."     Centralb.    f. 

Bakt.,  i  Abt,  Bd.  xx,  1896,  No.  10-11,  pp. 

393-395. 
KANTHACK,  A.   A.     Ueber  verzweigte   Diph- 

theriebacillen.     Centralb.    f,    Bkt,    xx    Bd., 

1896,  pp.  296-297. 

('96).  ZETTNOW.  Bilder  von  Spirillum  undula  majus 
bei  freiwilligem  Absterben.  Centralb.  f. 
Bakt.,  xix  Bd.,  1896,  pp.  177-180,  with  i 
plate,  16  figs. 

THAXTER,  ROLAND.  Contributions  from  the 
Cryptogamic  Laboratory  of  Harvard  Uni- 
versity, xxxix.  Further  Observations  on 
the  Myxobacteriaceae.  Botanical  Gazette, 
vol.  xxin.  No.  6,  1897,  pp.  393-411,  2  plates. 

('97)  MARPMANN,  G.  Zur  Morphologic  und  Biologic 
des  Tuberkelbacillus.  Centralb.  i.  Bakt., 
xxii  Bd.,  1897,  pp.  582-586,  with  i  plate. 


('96). 


('97). 


GENERAL    MORPHOLOGY   OF   THE    BACTERIA;    CYTOLOGY. 


2I7 


('97).  JOHAN-OLSEN,  OLAV.  Zur  Pleomorphismus- 
frage.  Centralb.  f.  Bakt.,  2  Abt.,  Bd.  in, 

1897,  pp.  273-284,  2  plates. 
Finds  branching  forms. 

('97).  KiTT.    Die  Streptothrixform  des  Rotlaufbacil- 

lus.     Centralb.  f.  Bakt.,  xxn  Bd.,  1897,  pp. 

726-732,  with  4  figs. 
('98).  SCHULTZ.    See  xxxvm. 
('98).  STOLZ,  ALBERT.    Ueber  besondere  Wachstums- 

formen    bei    Pneumo-    und    Streptokokken. 

Centralb.  f.  Bakt.,  xxiv  Bd.,  1898,  pp.  337- 

343,  with  6  figs. 

Figures  look  like  involution  forms. 

('98).  ZIEMANN,  HANS.  Eine  Methode  der  Doppel- 
farbung  bei  Flagellaten,  Pilzen,  Spirillen 
und  Bakterien,  sowie  bei  einigen  Amoben. 
Centralb.  f.  Bakt.,  xxiv  Bd.,  1898,  pp.  945- 
955,  i  plate. 

The  author's  figures  show  the  body  of  the  spirillum 
blue  enclosing  1-5  carmin  colored  granules. 

('98).  RUZICKA,  VLAD.  Zur  Frage  von  der  inneren 
Struktur  der  Mikroorganismen.  Centralb.  f. 
Bakt.,  xxin  Bd.,  1898,  pp.  305-307,  with  I 
plate. 

Kinds  granules,  which  take  stains,  in  bacterial  body 
reserves  conclusions  as  to  their  nature,  but  thinks  they 
are  not  pleomorphic  or  degeneration  phenomena. 

('98).  GRASSBERGER,  R.  Zur  Frage  der  Scheinfaden- 
'bildung  in  Influenzaculturen.  Centralb.  f. 
Bakt.,  xxin  Bd.,  1898,  pp.  353-364,  with  I 
plate  and  4  text  figs. 

('98).  BURCHARD,  GEORG.  Beitrage  ztir  Morphologic 
und  Entwickelungs-Geschichte  der  Bacte- 
rien.  Arb.  a.  d.  Bact.  Institut  der  tech. 
Hochschule  zu  Karlsruhe,  n  Bd.,  i  Heft, 

1898,  pp.  1-64,  2  pi. 

Twenty  new  species  are  described— Bacterium  pitui- 
taus,  B.  perittomaticum,  B.  flexile,  B.  turgescens,  B. 
bracliysporum,  B.  implectans,  B  petroselini,  B.  augulans, 
Bacillus  goniosporus,  B.  pectocutis,  B.  paucicutis,  B. 
cylindrosporus,  B.  leptodermis,  B.  bipolaris,  B.  loxo- 
sus,  B.  myxodens,  B.  armoraciae,  B.  idosus,  B.  loxosporus. 
B.  cursor. 

('98).  WAGNER,  A.  Coli-  und  Typhusbakterien  sind 
einkernige  Zellen.  Centralb.  f.  Bakt.,  xxm 
Bd.,  1898,  pp.  433-438,  and  pp.  489-492,  wifch 
2  plates  and  6  figs. 

('98).  CRAIG,  CHARLES  F.  The  branched  form  of  the 
bacillus  tuberculosis  in  sputum.  The  Journ. 
of  Exp.  Med.,  vol.  in,  1898,  pp.  363-370,  i 
plate. 

Author  thinks  it  is  premature  to  separate  B.  tubercu- 
losis from  the  bacteria  on  account  of  this  phenomenon. 

('99).  SCHULZE,    OTTO.      Unitersuchungen    iiber    die 

Strahlenpilzformen  des  Tuberculoseerregers. 

Zeitschr.  f.  Hyg.     Bd.  xxxi,  1899,  pp.  153- 

186,  i  plate. 
('99).  GALLI-VALERIO,  BRUNO.    Contribution  a  1'etude 

de     la     morphologic     du     Bacillus     mallei. 

Centralb.   f.  Bakt.,  xxvi  Bd.,   1899,  pp.   177- 

180,  with  5  figs. 
Author  finds  branched  forms  in  bouillon  and  on  agar. 

('99).  BERESTNEW,  N.  Zur  Frage  der  Klassifikation 
und  systematischen  Stellung  der  Strahlen- 
pilze.  Centralb.  f.  Bakt.,  xxvi,  1899,  p.  390. 

('99).  MOELLER,  ALFRED.  Ein  neuer  saure-  und  alko- 
holfester  Bacillus  aus  der  Tuberkelbacillen- 
gruppe,  welcher  echte  Verzweigungsformen 
bildet.  Centralb.  f.  Bakt.,  Bd.  xxv,  1899,  pp. 
369-373,  with  i  plate. 


('99).  SPIRIG,  W.  Die  Streptothrix  (Aotinomyces) 
Natur  des  Diphtheriebacillus.  Centralb.  f. 
Bakt,  xxvi  Bd.,  1899,  pp.  540-541. 

('99).  LUBARSCH,  O.  Zur  Kenntniss  der  Strahlen- 
pilze.  Zeitschr.  f.  Hyg.  Bd.  xxxi,  1899, 
pp.  187-220,  i  plate. 

('99).  MARX,  HUGO.  Zur  Morphologic  des  Rotz- 
bacillus.  Centralb.  f.  Bakt.,  xxv  Bd.,  1899, 
pp.  274-278,  with  4  figs.,  showing  branched 
forms. 

('99).  MUEHLSCHLEGEL,  A.  Ein  Beitrag  zur  Mor- 
phologie  und  Entwickelungsgeschichte  der 
Bakterien  nach  Studien  an  drei  Korner- 
baciHen.  Arb.  a.  d.  kaiserl.  Gesundheits- 
amte,  Bd.  xv,  Heft,  i,  pp.  131-152,  1899.  i 
plate  partly  colored.  Rev.  in  Centralb.  f. 
Bakt.,  xxv  Bd.,  1899,  p.  771. 
73  titles  cited  under  literature. 

Coo).  FEINBERG.  Ueber  den  Bau  der  Bakterien. 
Centralb.  f.  Bakt.,  xxvn  Bd.,  1900,  pp.  417- 
426,  with  5  plates. 

Author  believes  he  has  demonstrated  the  existence  of  a 
nucleus  in  the  bacteria  by  means  of  Romanowski's  stain- 
ing method  (a  mixture  of  methylene  blue  and  eosin).  The 
plasma  stains  blue  ;  the  nucleus,  which  may  be  small  or 
which  may  fill  nearly  the  whole  bacterial  body,  stains 
red  or  red-brown. 

Coo).  ZETTNOW.     Romanowski's    Farbung   bei    Bak- 
terien.    Centralb.  f.  Bakt.,  xxvn  Bd.,  1900, 
pp.  803-805. 
Says  Dr.  Feinberg's  papers  contribute  "  nidus  Neues  " 

Coo).  NAKANISHI,  K.  Vorlaufige  Mitteilungen  uber 
cine  neue  Farbungsmethode  zur  Darstellung 
des  feineren  Baues  der  Bakterien.  Munch, 
med.  Wochenschr.,  1900,  No.  6.  Rev.  in 
Centralb.  f.  Bakt.,  xxvn  Bd.,  1900,  pp.  547- 
549- 

"  All  bacteria  in  their  young  stage,  when  they  have 
grown  under  favorable  conditions,  are  one-nucleate 
short  cells." 

Coo).  SKSCHIVAN,  T.     Zur   Morphologic   des   Pest- 
bakteriutns.     Centralb.   f.  Bakt.,  xxvin  Bd., 
1900,  pp.  289-292,  with  4  text  figs. 
Finds  V-shaped  and  branched  forms. 

Coo).  MARX,  HUGO,  AND  WOITHE,  FRIEDRICH.  Mor- 
phologische  Untersuchungen  zur  Biologic 
der  Bakterien.  Centralb.  f.  Bakt.,  xxvin 
Bd.,  1900,  pp.  i-ii,  33-39,  65-69,  and  97-111, 
with  3  plates. 

Coo).  GALLI-VALERIO,  BRUNO.  Seconde  contribution 
a  1'etude  de  la  morphologic  du  B.  mallei. 
Centralb.  f.  Bakt.,  xxvm  Bd.,  1900,  pp.  353- 
359,  with  26  figs. 

Coi).  MEYER,  ARTHUR.  Ueber  die  Verzweigung  der 
Bakterien.  Centralb.  f.  Bakt.,  i  Abt.,  xxx 
Bd.,  1901,  No.  2,  pp.  49-60,  2  plates. 

Coi).  ROSENFELD.    See  xv. 

Coi).  REICHENBACH,    H.      Ueber   Verzweigung   bei 

Spirillen.     Centralb.    f.    Bakt.,    i    Abt.,   Bd. 

xxix,  1001,  pp.  553-557-     i  heliotype  plate. 

Many  of  the  spirilla  are  shown  with  distinct  branches. 

('02).  BUETSCHLI,  OTTO.    Bemerkungen  fiber  Cyano- 
pbyceen  und  Bacteriaceen.     Archiv.  f.  Pro- 
tistenkunde,  Bd.   i,  Heft.    i.     Jena,   Gustav 
Fischer,  1902.    pp.  41-58,  i  plate. 
On  the  nature  of  the  "  Centralkoerper." 

('02).  ERRERA,  L£o.  Sur  une  bacterie  de  grandes 
dimensions :  Spirillum  colossus.  Recueil  de 
1'Inst.  botanique  (Univ.de  Bruxelles),  T. 
v,  1902,  pp.  347-357-  Also  a  separate. 


218 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('02).  LOEB,  L.  M.     On  branching  forms  of  certain 
bacteria.     Jour,    of    Med.    Research.     Vol. 
vin,  1902  (n.  s.,  vol.  in),  pp.  415-423. 
The  author  finds  branching  forms  in  the  typhoid  ba- 
cillus. 

('02).  HEFFERAN,  MARY.  An  unusual  bacterial 
grouping.  Centralb.  f.  Bakt.,  2  Abt.,  vm 
Bd.,  1902,  pp.  689-699.  Also  a  separate, 
(with  5  figures  in  the  text). 

B.   rosaceus    metalloides  forms  rosette-like  groupings 
in  liquid  media  and  on  some  solid  media. 

('02).  MATZUSCHITA,  TEISI.  Beobachtungen  iiber 
den  merkwiirdigen  Teilungsprocess  bei  einem 
proteusartigen  I/uftbacillus.  (Vorl.  Mit- 
teilung).  Centralb.  f.  Bakt.,  Abt.  2,  Bd.  ix, 
1902,  pp.  257-260,  mil  I  Taf. 

('03).  HILL,  H.  W.,  AND  RICHARDS,  B.  R.  Notes  on 
Morphology,  i.  "Snapping"  as  character- 
istic of  the  diphtheroid  group,  n.  Branch- 
ing of  the  organism  of  actinomycosis.  Am. 
Pub.  Health  Ass'n.  Proc.  3(rth  Ann.  meet- 
ing -at  New  Orleans,  Dec.  9-12,  1902,  Co- 
lumbus, Ohio,  1903.  Also  a  separate,  4  pp., 
2  figs. 

('03).    WOLBACH,   S.    B.,   AND  ERNST,   HAROLD   C.      Ob- 

servations  on  the  morphology  of   Bacillus 
tuberculosis  from  human  and  bovine  sources. 
Jour,  of  Med.  Research,  1903,  vol.  x,  No.  3, 
PP-  3I3-333-     13  plates.    Also  a  separate. 
('04).  THAXTER,   ROLAND.      Contributions    from   the 
cryptogamic  laboratory  of  Harvard  Univer- 
sity, LVI,     Notes  on  the  Myxobacteriaceae. 
Bot.  Gaz.,  June,  1904,  vol.  xxxvii,  pp.  405- 
416,  with  2  plates. 
Cystobacter  becomes  Polyangium. 

('04).  LEPESCHKIN,  W.  W.  Zur  Kenntniss  der  Erb- 
lichkeit  bei  den  einzelligen  Organismen. 
Die  Verzweigung  und  Mycelbildung  bei 
einer  Bakterie  (Bacillus  Berestnewi  n.  sp.). 
Centralblatt  f.  Bakt.,  2  Abt.,  1904,  Bd.  xii, 
pp.  641-648,  and  Bd.  xni,  pp.  13-22,  20  figs. 
Describes  a  branched  form. 


XI.    Spores. 

[See  also  some  papers  under  X,  XV,  and  XXXIV.] 

('76).  COHN,  FERDINAND.     Untersuchungen  tiber  Bac- 

terien.     Beitrage  zur  Biologie  der  Pflanzen, 

Bd.  n,  Heft  2,  1876,  pp.  249-276. 
('84).  BUCHNER,  H.    Ueber  das  Verhalten  der  Spalt- 

pilz-Sporen  zu  den  Anilinfarbstoffen.  Aertz- 

liohe   Intelligenzbl.,   Jahrg.   xxxi,    1884,   pp. 

370. 
('87).  LEHMANN,  K.   B.     Ueber   die   Sporenbildung 

bei   Milzbrand.   Miinch.   med.   Wochenschr., 

1887,  pp.  485-488,  No.  26. 
('88).  PRAZMOWSKI,    A.     Ueber    Sporenbildung   bei 

den  Bakterien.     Verhand.  d.  k.  k.  Akad.  d. 

Wissenschaften  in  Krakau.     Math.-naturw. 

Sektion,  Bd.  xvm,  1888,  p.  35,  I  Tafel. 
Author   thinks   existence   of  "  arthrospores "    is    not 
proved. 

('88).  GLOBIG.    See  xxxm  and  xxxiv. 

('88).  GRUBER.    See  xxxm. 

('88).  BUCHNER,     H.       Ueber     die     vermeintlichen 

Sporen    der    Typhusbacillen.      Centralb.    f. 

Bakt.,  1888,  Bd.  iv,  pp.  385-390. 


('89).  KLEIN,  LUDWIG.  Ueber  einen  neuen  Typus 
der  Sporenbildung  bei  den  endosporen  Bac- 
terien.  Ber.  d.  deutsoh.  Botan.  Gesellsch., 
Bd.,  vn,  1889,  pp.  (57)-(72).  i  plate. 

Five  spore-bearing  bacilli  are  described.  The  spores 
are  green.  In  one  filamentous-septate  species  the  spores 
generally  occur  in  pairs,  with  a  septum  between,  i.  e.  ,  the 
spore  is  situated,  if  oue  may  so  speak,  in  the  positive 
pole  of  one  segment,  and  in  the  negative  pole  of  its  fel- 
low. 

('91).  FISCHER.    See  XL. 

('91).  MoELLER,  H.     Ueber  eine  neue  Methode   der 

Sporenfarbung.     Centralb.   f.   Bakt.,   Bd.   x, 

1891,  pp.  273-277- 

The  author  summarizes  his  method  as  follows  :  "  The 
air-dried  cover  glass  preparation  is  passed  through  the 
flame  three  times  and  put  into  absolute  alcohol  for  two 
minutes  ;  then  two  minutes  in  chloroform  ;  then  washed 
with  water  one-half  to  two  mimites  It  is  subsequently 
plunged  into  5  per  cent  chromic  acid  and  again  thor- 
oughly washed  in  water,  after  which  carbol-fuchsin  is 
dropped  upon  it  and  warmed  over  the  flame  to  boiling 
for  60  seconds.  The  carbol-fuchsin  is  then  poured  off,  the 
cover-glass  plunged  into  5  percent  sulphuric  acid  until 
it  is  bleached,  and  then  thoroughly  washed  in  water. 
The  watery  solution  of  methylen  blue  or  malachite  green 
is  then  allowed  to  act  upon  it  for  30  seconds,  after  which 
it  is  washed  ®ff.  The  spores  should  then  be  visible  as  a 
dark  red  in  a  beautiful  green  or  blue  bacterial  body." 

('91).  CRAMER,  E.  Die  Ursache  der  Resistenz  der 
Sporen  gegen  trockene  Hitze.  Arch.  f. 
Hygiene,  1891,  Bd.  xni,  pp.  71-112. 

Resistance  is  due  to  extreme  drvness  of  the  spores,  and 
to  the  fact  that  all  their  water  is  hygroscopic  water, 
which  evaporates  quickly  in  dry  air,  leaving  presumably 
pure  water-free  albumen. 

('92).  FOTH.  Zur  Frage  der  Sporenfarbung.  Cen- 
tralb. f.  Bakt.,  xi  Bd.,  1892,  pp.  272-278. 

Does  not  find  Moeller's  chromic  aciil  method  universally 
applicable.  Author  sometimes  substitutes  chloriodid 
zinc.  He  has  also  found  hydrogen  peroxide  very  useful. 
He  also  uses  aniliu  water  fuchsin.  A  pure,  colorless, 
toluidin  free  anilin  may  be  had  from  Jon.  Wolff,  in  Bres- 
lau.  In  case  oi  anthrax  spores,  the  proper  time  of  expos- 
ure is  i%  minutes  with  chromic  acid  ;  2  to  2%  minutes 
with  concentrated  chlorzinc  iodide  solution,  and  3  min- 
utes with  hydrogen  peroxide.  Here  also  the  author  says 
he  obtained  the  best  results  with  H2Oa. 

('93).  FIOCCA,  R.  Ueber  eine  neue  Methode  der 
Sporenfarbung.  Centralb.  f.  Bakt.,  xiv,  1893, 
No.  i,  pp.  8-9. 

Cover-glass  preparations  are  plunged  3  to  15  minutes 
(mostly  3  to  5  minutes)  into  steaming  hot  water  contain- 
ing 10  per  cent  ammonia  water,  to  which  has  also  been 
added  10  to  20  drapsof  some  alcoholic  anilin  solution. 
Covers  thus  treated  are  then  plunged  for  a  moment  into 
water  containing  20  per  cent  sulphuric  or  nitric  acid,  and 
are  subsequently  exposed  to  a  contrast  stain.  The  stains 
recommended  are  gentian  violet,  fuchsin,  methylene 
blue  and  safranin  ;  the  contrast  stains  are  vesuvin,  chry- 
soidin,  methylene  blue,  malachit  green  or  safrauin.  The 
preparations  are  said  to  be  clear  and  satisfactory. 

('93).  PHYSALIX.    See  xxxm. 

('94).  ERNST,  PAUL.  Farbungsversuche  an  Sporen 
mit  Hilfe  der  Maceration.  Centralb.  f.  Bakt., 
Bd.  xvi,  1894,  pp.  182-  184. 

('95).  MIQUEL.  P.,  AND  LATTRAYE,  E.  De  la  resis- 
tance des  spores  des  baoteries  aux  tempera- 
tures humides  egales  et  superieures  a  100 
degrees.  Ann.  de  micrographie.  Tome  vn, 
1895,  p.  no,  158,  205.  Rev.  in  Centralb.  f. 
Bakt.,  xix  Bd.,  1896,  pp.  360-362. 

('95).  BUNGE,  R.  Ueber  Sporenbildung  bei  Bakter- 
ien. Fortschr.  d.  Med.,  Bd.  xni,  1895,  No. 
20  and  21.  Rev.  in  Centralb.  f.  Bakt.,  xvm 
Bd.,  1895,  p.  718. 


SPORES ;  FLAGELLA. 


219 


('96).  SCHREIBER,  OSWALD.  Ueber  die  physiologis- 
chen  Bedingungen  der  endogenen  Sporen- 
bildung  bei  Bacillus  anthracis,  subtilis,  und 
tumesccns.  Centralb.  f.  Bakt.,  xx  Bd.,  1896, 
353-374  and  429-437. 
Forty-five  papers  cited  at  close  of  this  article. 

('96).  BUCHNER,  II.  Ueber  die  physiologischen  Be- 
dingungen der  Sporenbildung  beim  Milz- 
brandbacillus.  Centralb.  f.  Bakt.,  xx  Bd., 
1896,  pp.  806-807. 

("98).  CATTESINA,  G.  Ricerche  suH'intima  struttura 
delle  spore  dei  batteri.  Separate  from  Atti 
d.  Soc.  veneto-trentina,  vol.  in,  Fasc.  2,  Pa- 
dova,  1898,  10  pages,  with  I  plate.  Rev.  in 
Centralb.  f.  Bakt.,  xxvi  Bd.,  1899,  pp.  35-36. 

Some  evidence  in  favor  of  existence  of  a  nucleus. 

('98).  MIGULA,  W.  Der  Keimgehalt  und  die  Wider- 
standsfahigkeit  der  Bakterien  der  animalen 
Lymphe.  Arb.  a.  d.  Bact.  Institut  der  tech. 
llochschule  zu  Karlsruhe,  u  Bd.,  i  Heft, 

1898,  pp.  65-72. 

('98).  AUJESZKY,  ALADAR.  Eine  einfache  Sporen- 
farbungsmethode.  Centralb.  f.  Bakt.,  xxm 
Bd.,  1898,  pp.  329-33I- 

The  unfixed  covers  are  placed  in  a  boiling  hot  %  per 
cent  solution  of  HC1  for  3  to  4  minutes,  then  washed  in 
water,  dried,  fixed,  and  stained  with  hot  carbol  fuchsin 
(three  times  over  flame).  Covers  are  then  cooled,  bleached 
in  4  to  5  per  centsulphuric  acid,  and  couuterstaiued  I  to 
i  minutes  in  malachit  green  or  methylene  blue. 

('99).  STEPHANIDIS,  PHILOPIMIN.  Ueber  den  Einftuss 
des  NiihrstoffgehaJtes  von  Nahrboden  auf 
die  Raschheit  der  Sporenbildung  und  die 
Zabl  und  Resistenz  der  gebildeten  Sporen. 
Arch.  f.  Hyg.,  Bd.  xxxv,  1899,  PP-  r-10- 
Review  in  Centralb.  f.  Bakt.,  xxvi  Bd.,  1899, 
p.  568. 

In  a  poor  substratum  anthrax  spores  were  formed  more 
rapidly  but  in  less  numbers  than  in  a  rich  medium.  To- 
ward heat  the  spores  from  the  rich  and  poor  media  be- 
haved alike. 

('99).  KI,EIN,  ALEX.  Eiire  einfache  Methode  zur 
Sporen fartbung.  Cenitralb.  f.  Bakt.,  xxv  Bd., 

1899,  PP-  376-379- 

Klein's  modification  consists  in  staining  the  spores  be- 
fore they  have  dried.  In  a  watch  glass  he  makes  a  spore 
emulsion  in  phys.  salt  solution.  To  this  is  added  an 
equal  volume  of  filtered  carbol  fuchsin.  This  is  then 
gently  heated  over  the  open  flame  for  six  minutes,  i.  e. , 
until  steam  rises.  Covers  are  now  prepared  and  the  bac- 
terial layer  fixed  by  passing  twice  through  the  flame. 
The  covers  are  then  passed  through  I  per  cent  H2SO4  for 
1  to  2  seconds,  washed  in  water  and  counterstained  3  to  4 
minutes  in  alcoholic  methylene  blue  solution  diluted 
with  water. 

('99).  DANNAPPEI,.  See  xxxm. 
Coo).  SMITH,  R.  GREIG.  The  double  staining  of 
spores  and  bacilli.  Proceedings  of  the  Lin- 
nean  Society  of  New  South  Wales,  1900, 
Part  3,  June  27,  pp.  394-397.  Also  a  separate 
(issued  Nov.  22,  1900). 

('02).  SCHAUDINN,  FRITZ.  Beitrage  zur  Kenntnis 
der  Bakterien  und  verwandter  Organismen. 
i.  Bacillus  biitchlii,  Arch.  f.  Protistenkunde, 
Bd.  i,  1902,  pp.  306-343,  i  plate.  Bibliography 
of  24  titles. 

This  very  large,  slow-moving  organism  was  isolated 
from  the  intestinal  tract  of  a  cockroach,  Periplaneta 
orientalis. 

The  author  states  that  this  organism,  like  Kern's  Di- 
spora  caucasica,  is  constantly  disporous.  B.  buetchlii  was 
selected  for  study  of  its  inner  structure,  on  account  of  Its 
large  size.  The  membrane  did  not  give  the  cellulose  re- 
action. 


Seventy-three  figures  are  (*iven(  illustrating  inner 
structure,  stages  in  the  formation  of  the  spores  (one  in 
each  pole  ofthe  rod),  polar  germination  of  the  spores,  etc. 
The  organism  is  24  to  80  ^  long  by  3  to  6  n  broad,  mostly 
50  to  60  (i  x  4  to  5  (».  The  bacillus  is  flagellate,  after  the 
manner  of  B.  subtilis. 


XII.     Flagella. 

('38).  EHRENBERG.    See  v. 
('72).  COHN.    See  v. 

('75).  DALLINGER,  W.  H.,  AND  DRYSDALE,  J.  J.  On 
the  existence  of  flagella  in  Bacterium  termo. 
The  Monthly  Microscopical  Journal,  Sept. 
I,  1875,  PP-  105-108. 

('76).  WARMING,  EUG.  Om  nogle  ved  Danmarks 
Kyster  levende  Bakterier.  Kjobenhavn, 
1876. 

('77).  KOCH.    See  LV. 

('78).  DALUNGER,  W.  H.  On  the  measurement  of 
the  diameter  of  die  flagella  of  Bacterium 
termo :  a  contribution  to  the  question  of  the 
"Ultimate  limit  of  vision"  with  our  present 
lenses.  Journ.  Roy.  Micros.  Soc.,  vol.  i, 
1878,  pp.  169-175,  2  plates. 

From  the  mean  value  of  200  measurements  (50  with 
each  of  4  high-power  objectives)  Dallinger  concludes  that 
the  diameter  of  the  unstained  flagellum  of  B.  termo,  in 
round  numbers,  is  one-two  hundred  and  four  thousandth 
(1-204000)  of  an  inch.  This  is  equal  to  about  one-eighth 
micron. 

('79).  VAN  TIEGHEM,  PHILIPPE  EDOUARD  L£ON.  Sur 
les  pretendus  cils  des  bacteries.  Bull,  de  la 
Societe  Botanique  de  France,  1879,  T.  xxvi, 
PP-  37-45- 

Van  Tieghem  maintained  that  the  flagella  were  moved 
from  within  the  body  of  the  bacterium,  they  themselves 
being  inert  gelatinous  organs,  and  not  vibratile  cilia. 

('89).  LOEFI'LER,  F.  Eine  neue  Methode  zum  Farben 
der  Mikroorganismen,  im  besonderen  ihrer 
Wimperhaare  und  Geisseln.  Centralb.  f. 
Bakt.,  vi  Bd.,  1889,  No.  8-9,  pp.  209-224,  mit 
8  Photogrammen. 

('89).  TRENKMANN.     Die  Farbung  der  Geisseln  von 
Spirillen  und   Bacillen.     Centralb.   f.   Bakt., 
vi  Bd.,  Oct.  15,  1889,  No.  16-17,  pp.  433-436. 
('90).  MESSEA.    See  LVI. 

('90).  LOEFFLER,  F.  Weitere  Untersuchungen  fiber 
die  Beizung  und  Farbung  der  Geisseln  bei 
den  Bakterien.  Centralb.  f.  Bakt.,  Bd.  vn, 
1890,  pp.  625-639. 

('90).  TRENKMANN.     Die  Farbung  der  Geisseln  von 

Spirillen  und  Bacillen.    n.  Mitth.    Centralb. 

f.  Bakt.,  1890,  Bd.  vm,  No.  13,  pp.  385-389. 

Covers  on  which  the  bacterial  film  is  dried  without  heat 

are  put  for  6  to  12  hours  In  water  containing  2  per  cent 

tannin  and  0.5  to  0.25  of  one  per  cent  hydrochloric  acid. 

They  are  subsequently  washed  for  one  hour  in   iodine 

water,  and  then  stained  %  hour  in  weak  gentian  violet 

anilin  water,  made  as  follows  :    Into  a  test  tube  holding 

25  cc.  put  a  few  drops  of  concentrated  alcoholic  solution 

of  gentian  violet  and  add  10  cc.  of  distilled  water.    Then 

pour  out  about  one-half  of  this  and  fill  up  with  anilin 

water.   The  clear  stain  is  said  to  color  the  flagella  well  on 

a  feebly-stained  background. 

('91).  HUMPHREY,  J.  E.     Notes  on  Technique,     n. 

Rot.  Gazette,  1891,  pp.  71-73. 

Cilia  of  zoospores  of  alga;  and  fungi  are  stained  very 
readily  and  sharply  "in  a  drop  of  moderately  strong  solu- 
tion (in  qo  per  cent  alcohol)  of  Hanstein's  rosanilliu- 
violet,  composed  of  equal  parts  of  fuchsin  and  methyl 
violet,"  after  first  fixing  them  in  a  couple  of  drops  of  i 
per  cent  osmic  acid  solution. 


22O 


BACTERIA   IN    RELATION    TO    PLANT   DISEASES. 


('92).  STRAUSS,  I.  Sur  un  precede  de  coloration  a 
1'etat  vivant  des  cils  ou  flagella  de  certiaines 
bacteries  mobiles.  C.  R.  de  la  Soc.  de 
biologic,  1892,  No.  23,  pp.  S42-S43.  Also 
Bull.  Med.,  1892,  p.  1,003. 

('93).  SO.AVO.     Di  un  rapido  processo  per  le  colora- 
zione  delle  ciglia  di  alcuni  microorganism!. 
Ministera  dell'intern.     Laboraitori  scientifica 
della  direzione  di  Santa-Roma. 
Rev.  in  Centralb.  f.  Bakt.  Bd.  xv.  p.  507, 1893. 

('93).  NICOLLE,  M.,  ET  MORAX,  V.  Technique  de  la 
coloration  des  cils,  etc.  Ann.  de  1'Inst.  Pas- 
teur, T.  vii,  1893,  No.  7,  pp.  554-56i. 

('93).  MOORE,  V.  A.  On  the  character  of  flagella  on 
the  Bacillus  choleraesuis,  B.  coli  communis, 
and  the  B.typhi  abdominalis.  Wilder  Quar- 
ter Century  Book,  Ithaca,  N.  Y.,  1893,  pp. 
339-363. 

('93).  VAN  ERMENGEM,  E.  Nouvelle  methode  de 
coloration  des  cils  des  bacteries.  Trav.  du 
Lab.  d.  Hygiene  et  de  Bact.  de  1'Univ.  de 
Gand,  T.  I.,  1.  3,  1893- 

Original  not  seen.  A  very  good  method.  Reviewed  in 
Zeitschr.  f.  Wissensch.  Mikr.  Bd.  xi.  1894,  pp.  98-99,  and 
in  Ann.  de  Micrographie,  T.  V.,  1893,  pp.  394-395- 

('94).  HESSERT,  W.  Geisselfarbung  ohne  Beize. 
Centralb.  f.  Bakt.,  Bd.  xvi,  1894,  No.  8-9, 

PP.  346-347- 

See  also  A  simple  stain  for  ciliated  bacteria.  Chicago 
Med.  Recorder,  1894,  pp.  240-242. 

('94).  BUNGE,   R.     Ueber   Geisselfarbung  von   Bak- 
terien.    Fortschr.  d.  Medizin,  Bd.  xn,  1894, 
No.  12,  pp.  462-464. 
('94).  BUNGE,  R.     Zur  Kenntniss  der  Geisseltragen- 

den  Bakterien.    Ibid.,  No.  17,  pp.  653-670. 
('94).  BUNGE,  R.    Weitere  Mittheilungen  iiber  Geis- 
selfarbung.    Fortschr.    d.    Mediz.,    Bd.    xn, 
1894,  pp.  929-935- 

('95).  MOORE,  VERANUS  A.  On  ithe  nature  of  the 
flagella  and  their  value  in  the  systematic 
classification  of  the  bacteria.  Jour.  Am. 
Pub.  Health  Asso.,  Oct.,  1895,  Ann.  vol.  xx, 
pp.  432-444,  3  plates. 

('95).  FERRIER.  Considerations  generates  sur  le 
pleomorphisme  des  cils  viforatiles  de  quelques 
bacteries  mobiles. — Archives  de  medicine 
experimentale  et  d'anatomie  pathologique. 
Paris,  Serie  I.  T.  vn,  1895,  pp.  58-75-  I 
plate. 

('96).  LOEWIT,  N.  Zur  Morphologie  der  Bakterien. 
Centralb.  f.  Bakt.,  xix  Bd.,  1896,  pp.  673- 
686,  with  i  plate. 

('96).  KANTHACK,  A.  A.,  AND  CONNELL,  T.  W.  The 
flagella  of  the  tetanus  bacillus,  and  other 
contributions  to  the  morphology  of  the 
tetanus  bacillus.  Jour.  Path,  and  Bact.,  iv, 
1896-97,  pp.  452-459- 

('98).  BOWHIIJ,,  TH.  Eine  neue  Methode  der  Bak- 
teriien-Geisselfarbung  bei  Gebrauch  einer 
Oroeinbedze.  Hyg.  Rundschau,  1898,  No.  I. 
Rev.  in  Centralb.  f.  Bakt.,  xxm  Bd.,  1898, 
pp.  667-668. 

('98).  STEPHENS,  J.  W.  Van  Ermengem's  method  of 
staining  flagella;  a  modification.  The  Lan- 
cet, 1898,  Oct.  i.  Rev.  in  Centralb.  f.  Bakt., 
xxv  Bd.,  1899,  p.  392. 

Substitutes  for  nitrate  of  silver  a  2  percent  largin  solu- 
tion. The  flagella;  are  said  to  be  cleaner  and  more  dis- 
tinct. 


('99).  WELCKE,  E.    Eine  neue  Methode  der  Geissel- 
farbung.   Arch.  f.  klin.  Chir.,  Bd.  ux,  1899, 
Heft,    i,  pp.    129-143.     Rev.   in   Centralb.   f. 
Bakt.,  Bd.  xxvi,  1899,  pp.  520-521. 
A  silver  process  of  about  the  same  complexity  as  that 
of  van  Ertnengem. 

('99).  ZETTNOW.  Ueber  Geisselfanbung  bei  Bakter- 
ien. Zeitschr.  f.  Hyg.,  Bd.  xxx.  i  Heft., 
March,  1899,  pp.  95-106. 

Discusses  van  Ermengem's  silver  method,  and  gives  a 
gold  method  which  is  said  to  be  better. 

('99).  MORTON,  N.  Flagella  staining.  Trans.  Jenner 
Inst,  London,  vol.  i.  2  series,  pp.  242-243, 
1899. 

Tap  water  is  recommended  for  dilutions.  A  24-hour 
agar  surface  growth  furnishes  the  bacteria.  They  are 
allowed  to  diffuse  in  a  little  of  this  sterilized  water  in  a 
watch  glass.  A  drop  or  two  of  this  is  then  placed  on  the 
clean  slide  or  cover  and  spread  as  widely  as  possible  with- 
out use  of  the  needle.  The  excess  is  absorbed  by  blotting 
paper.  The  dried  film  is  not  fixed  by  heat.  The  stain 
consists  of  taunic  acid  i  gram,  potash  alum  i  gram,  dis- 
tilled water  40  cc.  To  this  is  added  0.5  gram  of  night  blue 
dissolved  in  20  cc.  of  absolute  alcohol.  The  copious  pre- 
cipitate which  results  is  carefully  removed  by  filtration. 
The  fluid  is  then  ready  for  use.  Stain  2  minutes.  The 
flagella  are  blue,  the  body  of  the  organism  is  not  stained. 
Longer  exposures  cause  precipitates.  Couuterstaiu  for 
the  body  by  exposure  for  i  or  2  minutes  to  anilin-water 
gentian  violet.  "  I  consider  that  the  application  of  heat 
and  the  spreading  of  the  film  with  the  needle  are  very 
fatal  to  good  results  in  flagella  staining." 

('oo).  HINTERBERGER,  A.  Eine  Modifikation  des 
Geisselfarbungsverfahrens  nach  van  Ermen- 
gem.  Centralb.  f.  Bakt.,  Bd.  xxvn,  No. 
16-17,  I9°°.  PP-  597-605,  I  plate  and  I  fig. 

('oi).  WH.UAMS,  HUGH.  Flagella  stain.  See  Path- 
ological technique,  by  Mallory  and  Wright. 
Second  Ed.,  1901.  pp.  104-106.  W.  B. 
Saunders  &  Co.,  Philadelphia  and  London. 

('02).  MEYER,  ARTHUR.  Kurze  Mitteilung  iiber  die 
Begeisselung  der  Bakterien.  Centralb.  f. 
Bakt.,  Abt.  i,  Bd.  xxxi,  Originate,  1902, 
PP-  737-739- 


XIII.    Capsules. 

('78).  CIENKOWSKI.     Untersuohung  iiber  die  Gallert- 
bildungen   des   Zuckerriibensaftes.     Resume 
allemand  du  memoire  <russe,  Charkow,  1878. 
Not  seen. 

('85).  RiBBERT.    Zur  Farbung  der  Pneumoniekokken. 
Deutsche  med.  Wochenschr.,  11  Jahrg.,  1885, 
p.  136. 
Gives  a  method  for  staining  capsules. 

('85).  FRIEDI.AENDER,  C.  Ueber  Pneumonie-Micrococ- 
cen.  iFortsoh.  d.  Med.,  Bd.  in,  1885,  pp. 

91-93- 

Replies  to  criticism  of  Germain  See,  and  gives  his 
method  of  staining  the  capsule. 

('85).  FRIEDLAENDER,  C.  Notiz,  die  Farbung  der  Kap- 
selmicrococcen  betreffend.  Fortsch.  d. 
Medicin,  Bd.  in,  1885,  No.  23,  pp.  757-76o. 

The  author's  plan  for  staining  capsules  is  as  follows  : 
Pass  the  dried  covers  three  times  through  the  flame,  and 
then  expose  from  one  to  several  minutes  in  i  per  cent 
acetic  acid  water.  Remove  from  the  cover  by  blowing 
through  a  glass  tube  drawn  to  a  point,  and  dry  in  air 
quickly.  Then  expose  in  a  saturated  solution  of  anilin- 
water  gentian  violet  for  a  very  few  seconds,  wash  in  water 
and  examine.  The  exposure  to  the  gentian  violet  should 
be  barely  long  enough  to  stain  the  capsule  without  stain- 
ing the  interior  protoplasm. 


CAPSULES;  STAINS  AND  STAINING  METHODS. 


221 


('92).  LIESENBERG,  C.,  u.  ZOPF,  W.  Ueber  den  sogen- 
annten Froschlaichpilz  (Leuconostoc)  der 
europaischen  Ritbenzucker-  und  der  javan- 
ischen  Rohrzuckerfabriken.  Zopf  s  Beitraege, 
Hft.  I,  1892,  pp.  1-29,  with  2  plates,  and 
Hft.  ii,  1892,  pp.  1-2.  Rev.  in  Am.  Nat., 
March,  1897,  p.  228. 

('92).  WELCH,  WM.  H.  A  gas-producing  bacillus 
(B.  aerogenes  capsulatus,  nov.  spec.)  capable 
of  rapid  development  -in  the  blood-vessels 
after  death.  Bull.  Johns  Hopkins  Hospital, 
vol.  3,  No.  24,  July-August,  1892,  pp.  81-91. 
Also  a  separate. 

('94).  KOCH,  ALFRED,  AND  HOSAEUS,  HANS.  Ueber 
einen  neuen  Froschlaich  der  Zuckerfabriken. 
Centralb.  f.  Bakt,  Bd.  xvi,  1894,  pp.  225- 
228,  mit  i  fig. 

Describes  a  branching  gelatinous  organism  as  Bact. 
pedlculatum.  The  branching  is  due  to  a  terminal  split- 
ting or  one-sided  development  of  the  capsule. 

('95).  STIFT,  A.  Ueber  die  in  den  Produkten  der 
Zuckerfabrikation  auftretenden  Bakterien. 
Centralb.  f.  Bakt.,  2  Abt.,  Bd.  i,  1895,  pp. 
277-283. 

('96).  WILDE,  MAX.  Ueber  den  Bacillus  pneumonia: 
Friedlander's  und  verwandte  Bakterien. 
Inaugural  Dissertation,  Bonn,  1896,  pp.  74. 
Bibliography  of  89  titles.  Printed  by  Carl 
Georgi. 

('96).  FRICKE,  CARL.  Ueber  den  sogenannten  Bacil- 
lus mucosus  capsulatus.  Zeitschr.  f.  Hyg., 
Bd.  xxm,  1896,  pp.  380-451,  with  a  bibliog- 
raphy of  25  titles. 

('98).  PANE,  N.  Ueber  die  Genesis  der  Kapseln  des 
Pneumococcus.  Centralb.  f.  Bakt.,  xxiv  Bd., 

1898,  pp.  289-294,  with  21  figs. 

The  capsule  is  the  swollen  outer  part  of  the  bacterium. 
('98).  KAUFMANN.    Eine  neue  Methode  zur  Farbung 
von    Bakterienkapseln.      Hyg.    Rundschau, 
1808,   No.   18.     Abstr.   Centralb.   f.   Bakt.,   i 
Abt.,  Bd.  xxv,  .p.  32. 
('99).  WARD.    See  XLVII. 
('99).  MOORE,  A.     Capsule  staining.     Trans.  Jenner 

Inst.,  2d  ser.,  pp.  244,  1899. 

Recommends  a  contrast  stain  for  the  capsule.  The 
preparation  is  first  fixed  with  dilute  acetic  acid,  then 
stained  in  carbol  fnchsin  for  about  i  minute,  washed  in 
distilled  water  and  dried.  It  is  then  stained,  with  or 
without  gentle  heat,  in  night  blue  for  I  or  2  minutes, 
washed  and  dried.  The  night  blue  solution  is  Morton's 
modification  of  McCrorie's  stain.  (See  Morton,  Flagella 
staining.)  Hy  this  method  the  capsule  is  stained  blue  and 
the  body  dark  red. 

('99).  STRONG,  LAWRENCE  WATSON.  Ueber  die  Kap- 
selbacillen.  Centralb.  f.  Bakt.,  xxv  Bd., 

1899,  pp.  49-52. 

('oo).  BONI,  ICILIO.  Methode  zur  Darstellung  einer 
"  Kapsel "  bei  alien  Bakterienarten.  Centralb. 
f.  Bakt.,  xxviii  Bd.,  1900,  pp.  705-707. 

Coi).  WARD.    See  XLVII. 

('oi).  BONI,  I.  Ricerche  sulla  capsula  dei  batteri. 
Giorn.  Soc.  ital.  igiene,  Milano,  vol  xxm, 
1901,  pp.  417-430. 

('02).  SMITH,  R.  GREIG.  An  Ascobacterium  from 
the  sugar-cane,  with  notes  upon  the  nature 
of  the  slime  (Bacterium  saoohari,  n.  sp.). 
Proceedings  of  Linnean  Soc.  of  New  South 
Wales,  vol.  xxvn,  1902,  part  i,  pp.  137-145. 
i  plate.  Also  a  separate  (issued  Aug.  22, 
1902).  See  also  Centralb.  f.  Bakt.,  2  Abt, 
Bd.  ix,  p.  806. 


XIV.    Stains  and  Staining  Methods. 

(Sec  also  XI,  XII,  and  XIII.) 

(' — )•  EHRLICH.  Beitrage  zur  Kenntniss  der  Anilin- 
farbungen  und  ihrer  Verwendung  in  der 
mikroskopischen  Technik.  Arch.  f.  mikr. 
Anat.  Bd.  xm,  p.  263. 

('75).  WEIGERT.    Farbung  von  Bakterien.     Ber.  tiber 
d.    Sitzungen    d.    schlesischen    Gesellsch.    f. 
vatenl.  Cultur,  10  Dec.,  1875 
('77).  KOCH.    See  LV. 

('81).  WEIGERT.  Zur  teohnik  der  mikroskop.  Bak- 
terienuntersuchungen.  Virch.  Archiv.  Bd. 
LXXXIV,  1881,  p.  275. 

('82).  EHRLICH.  FaPbung  der  Tuberkelbacillen 
Verhdlgn.  d.  Ver.  f.  i.  Med.,  i  Mai,  1882 
D.  med.  Woch.,  1882,  p.  269. 

('82).  ZIEHL.  Zur  Farbung  des  Tuberkelbacillus. 
Deutsche  med.  Wochenschr.,  1882,  No.  33, 
P-  451. 

('82).  SZYSZYLOWICZ,  J.  Das  Corallin  als  mikro- 
chemisches  Reagens  in  d.  Pflanzenhistologie, 
R.  i.  S.  Ak.  Krakau,  T.  x,  1883,  pp.  97-114. 
Not  seen.  Rev.  Bot.  Centralb.,  1886,  Bd. 
xxvm,  p.  51.  See  also  Bot.  Centralb.,  Bd. 
xn,  1882,  p.  139. 

This  substance  should  be  dissolved  in  sodium  carbon- 
ate. It  is  used  as  a  test  for  slime  derived  from  starch 
which  stains  diffusely  purple  or  rose.  After  staining  the 
preparation  should  be  subjected  to  hot  alcohol,  which 
removes  the  stain  from  all  but  the  starch  grains  and 
shine  derived  therefrom.  Gum  slime  and  cellulose  slime 
are  bleached,  the  former  even  in  cold  alcohol.  Pure  gum 
does  not  stain .  Mount  the  preparation  in  Canada  balsam. 

('83).  ZIEHI,.  Zur  Lehre  von  den  Tuberkelbacillen, 
insbesondere  iiber  deren  Bedeutung  fiir 
Diagnose  und  Prognose.  Deutsche  med. 
Woohenschr.,  1883,  p.  62 

('84).  KOCH.    See  vi. 

('84).  LOEFFLER.    See  vi. 

('84).  GRAM,  C  Ueber  die  isolirte  Farbung  der 
Schizomyceten  in  Schnitt-und  Trocken- 
praparaten.  Fortsohr.  d.  Med.,  Bd  n  No 
6,  1884,  pp.  185-189. 

('86).  PFEFFER,  W.  Ueber  Aufnahme  von  Anilin- 
far.ben  in  lebende  Zellen.  Untersuchungen 
a.  d.  bot.  Inst.  Tubingen,  n,  p.  179,  1886. 

('87).  UNNA.  Die  Rosaniline  und  Pararosaniline, 
eine  bacteriologische  Farbenstudie.  Derma- 
tologischen  Studien.  Heft  4,  pp.  9-73. 
Hamburg,  Voss,  1887. 

('87).  KUEHNE.  Ueber  ein  combinirtes  Universal- 
verfahren,  Spaltpilze  im  thierischen  Gewebe 
naohzuweisen.  Dermatol.  Studien,  heraus- 
geg.  von  Unna.  Heft  6.  Hamburg,  L.  Voss, 
1887. 

('87).  HEIM,  L.     Die  Neuerungen  auf  dem  Gebiete 
der    bakteriologischen    Untersuchungsmetli- 
oden   seit    dem    Jahre,    1887,    I.    Farbungs- 
methoden.     Centralb.  f.  Bakt.,  x  Bd.,  1891, 
pp.  260-265,  288-296,  and  323-328. 
('88).  KUEHNE,  H.     Praktische  Anleitung  zum  mikro- 
skopischen    Naohweis     der     Bakterien     im 
thierisohen    Gewebe.      Leipzig    (Giinthers), 
1888,  pp.  15-23,  Methylenblaumethode. 
('88).  UNNA,    P.    G.      Die    Entwicklung    der    Bak- 
terienfarbung.        Eine      historisch-kritische 
Uebersioht.    Centralb.  f.  Bakt.,  1888,  in  Bd., 
pp.  22-26;  also  pp.  61-63,  93-99,  120-125,  153- 
158,   189-195,  218-221,  254-259,  285-291,  312- 
.    320,  345-348. 
At  the  end  71  papers  on  this  subject  are  mentioned  by 


222 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('91).  PREGL,  FRITZ.  Ueber  eine  neue  Karbolmethy- 
lenblau-Method.  Centralb.  f.  Bakt.,  x  Bd., 
1891,  pp.  826-829. 

Pregl  finds  the  following  method  of  staining  sections 
on  slides  satisfactory :  "  Exposure  for  one-half  to  one 
minute  to  carbol  methyleu  blue,  sometimes  with  the  aid 
of  heat;  second,  brief  washing  in  water  ;  third,  bleaching 
in  50  per  cent  alcohol  until  the  section  has  become  pale 
blue  passing  into  greenish  ;  fourth,  removal  of  the  water 
by  means  of  absolute  alcohol;  fifth,  clearing  in  xylol ; 
sixth,  enclosure  in  balsam." 

Kuehne's  carbol  methylen  blue  is  made  as  follows  :  1.5 
grins,  methylen  blue,  10  grms.  absolute  alcohol,  loo  grams  5 
per  cent  carbolic-acid  water.  The  alcohol  is  poured  over 
the  methylen  blue  •  the  carbolated  water  is  then  added  and 
rubbed  up  thoroughly  with  it.  If  only  a  small  quantity 
is  needed  from  time  to  time,  it  is  better  to  make  it  up  in 
less  amounts,  as,  with  time,  the  staining  power  of  the 
solution  decreases. 

('92).  KUEHNE,  H.  Das  Malachitgriin  als  Auszieh- 
ungsfarbe.  Centralb.  f.  Bakt,  xi  Bd.,  1892, 
pp.  756-758. 

Author  finds  malachit  green  dissolved  in  anilin  oil  very 
useful  for  treatment  of  sections  stained  in  carbol-fuch- 
sin,  etc.  The  bacteria  stand  out  distinctly  deep  red  on  a 
bluish  background.  See  paper  for  details. 

('93).  NICOLLE,  ET  CANTACUZENE,  J.  Proprietes 
colorantes  de  1'oxychlorure  de  ruthenium 
ammoniacal.  Ann.  de  1'Inst.  Pasteur,  T. 

vn,  pp.  331-334- 

('95).  UNNA,  P.  G.  Ueber  Verwendung  von  Anihn- 
mischungen  zur  tinktoriellen  Isolierung  von 
Gewebselementen.  Monatshefte  f.  prakt. 
Dermatologie,  Bd.  xxi,  1895.  Rev.  in  Cen- 
tralib.  f.  Bakt.,  I  Abt.,  xx  Bd.,  1896,  p.  406. 

('95).  NICOLLE,  M.  Pratique  des  colorations  micro- 
biennes,  Methode  de  Gram  modinee  et 
methode  directe.  Ann.  de  1'Inst.  Pasteur, 
T.  ix,  1895,  No.  9.  Rev.  in  Centralb.  f. 
Bakt,  xvni  Bd.,  1895,  p.  552. 

Carbol  violet  is  substituted  for  anilin  violet.  Excess  of 
color  is  removed  withalcohol-aceton. 

('97).  SEMENOWICZ,  W.,  UND  MARZINOWSKY,  E. 
Ueber  ein  besonderes  Verfahren  zur  Far- 
bung  der  Bakterien  im  Deckglaspraparate 
iund  in  Schnitten.  Centralb.  f.  Bakt,  xxi 
Bd.,  1897,  pp.  874-876. 

('oo).  DREYER,  GEORGES.  Bakterienfarbung  in  gleich- 
zeitig  nach  van  Gieson's  Methode  behandel- 
iten  Schnitten.  Centralb.  f.  Bakt.,  I  Abt., 
xxvn  Bd.,  1900,  pp.  534-535- 

Can  be  used  for  differential  staining  of  animal  tissues 
with  bacteria  in  them,  provided  the  latter  are  such  as 
stain  by  Gram's  method. 

('oo).  LE  Doux.     Bemerkungen  zu  dem  Artikel  des 
•   Herrn  M.  Dorset :  "A  new  stain  for  Bacillus 
tuberculosis."     Centralb.    f.    Bakt,    I    Abt, 
xxvn  Bd.,  1900,  p.  616. 

Dorsett  recommended  Sudan  III  for  staining  the  B. 
tuberculosis.  Le  Doux  says  he  has  tried  this  repeatedly 
with  entirely  negative  results. 

('02).  LEVINSON,  JA.  B.  Ueber  Farbung  des  Fettes 
und  der  fettigen  Degeneration  der  geform- 
ten  Elemente  in  flussigen  und  halbflussigen 
Medien  mit  Sudan  in.  Wratsch.  St. 
Petersburg,  1902,  Bd.  I,  pp.  1,208-1,209. 
(Russian.) 

('02).  GRIM  ME,  ARNOLD.  Die  wichtigsten  Methoden 
der  Bakterienfarbung  in  ihrer  Wirkung  auf 
die  Membran,  den  Protoplasten  und  die 
Einschliisse  der  Bakterienzelle.  Centralb.  f. 
Bakt.,  Abt.  I,  Bd.  xxxir,  Originale,  1902,  pp. 
1-16,  pp.  81-00,  pp.  161-180,  pp.  241-255,  pp. 
321-327,  with  2  plates.  Bibliog.  of  78  titles. 


XV.     Morphological  and  Physiological  Changes  Due 
to  Changed  Environment. 

('87).  SCHOTTEUUS.  Untersuchungen  iiber  den 
Microc.  prodigiosus.  Festsehr.  f.  Kolliker, 
Leipzig,  1887.  Not  seen. 

This  author  obtained  pigmeuted   and    uon-pigmented 
races  of  Bacillus  prodigiosus. 

('89).  BEHRING.  Beitriige  zur  Aetiologie  des  Milz- 
brandes.  vi.  Ueber  asporogenen  Milzbrand. 
Zeitschr.  i.  Hyg.,  1889,  Bd.  vn,  pp.  171-176. 

Author  describes  method  of  obtaining  Aplanobacter 
(Bacillus)  authracis  without  spores. 

('90).  SMITH.    See  vi. 

('90).  Roux,  E.  Baateridie  charbonneuse  asporogene. 
Ann.  de  1'Inst.  Pasteur,  T.  iv,  1890,  pp.  25-34. 

('92).  NENCKI,  M.  Ueber  Mischkulturen.  Cenfcralib. 
f.  Bakt.,  xi  Bd.,  1892,  pp.  225-228. 

('92).  ADAMI,  J.  G.  On  the  variability  of  bacteria 
and  the  development  of  races.  Med.  Chron- 
icle, vol.  xvi,  1892,  No.  6,  pp.  366-389.  Also 
a  reprint  from  Med.  Chronicle,  Sept.,  1892, 
8vo.,  26  pp.,  Manchester  (John  Heywood), 
1892. 

('92).  CHARRIN,  A.,  ET  PHISALIX.  Abolition  nersis- 
tante  de  la  fonction  chromogene  du  Bacillus 
pypcyaneus. — C.  R.  des  se.  de  1'Acad.  des 
Sci.,  Paris,  1892.  T.  cxiv,  pp.  1,565-1,568. 

(*93)-  SANDER.  Ueber  das  Wachstum  von  Tuber- 
kelbacillen  auf  pflanzlichen  Niihrboden. 
Arch.  f.  Hyg.,  Bd.  xvi,  Heft  3,  1893,  pp. 
238-311. 

At  38°-39°  C.  (not  at  22-23°  C.),  this  author  induced  the 
organism  of  tuberculosis  to  grow  on  a  variety  of  vegeta- 
ble substances,  i.e. .potato,  carrot,  kohlrabi,  macaroni, 
etc.  All  were  cooked. 

('93).  STONEY,  G.  Suggestion  as  to  a  possible  source 
of  the  energy  required  for  the  life  of  bacilli, 
and  as  to  the  cause  of  their  small  size. 
Proc.  Roy.  Soc.,  Dublin.  Vol.  viir,  Pt.  I, 
pp.  154-156.  Dublin,  1893. 
Views  of  a  physicist. 

('94).  SURMONT,  H.,  ET  ARNOULD,  E.  Recherohes 
sur  la  production  du  bacille  du  charbon 
asporogene.  Ann.  de  1'Inst.  Pasteur,  T.  vm, 
1894,  pp.  817-832. 

('94).  URY,  JAKOB.  U_eber  die  Schwankungen  des 
Bacterium  coli  commune  in  morphologis- 
cher  und  kultureller  Beziehung.  (Inaug. 
Diss.)  8vo.,  47  pp.,  Strassburg  i.  E.,  1894. 
Rev.  in  Centralb.  f.  Bakt.,  Bd.  xvi,  1894,  pp. 
579-58i. 

('94).  SMITH,  THEOBALD.  Modification,  temporary 
and  permanent,  of  the  physiological  char- 
acters of  bacteria  in  mixed  cultures.  Trans. 
Assoc.  Amer.  Physicians,  Phila.,  1894,  pp. 
85-109. 

('94).  DIEUDONNE,  A.  Beitrage  zur  Kenntnis  der 
Anpassungsfahigkeit  der  Bakterien  an  ur- 
sprunglich  ungiinstige  Temperaturverhalt- 
nisse.  Arbeiten  aus  dem  kaisenl.  Gesund- 
heitsamte,  Bd.  ix,  1894,  pp.  492-508,  Berlin, 
1894  (Julius  Springer).  Also  a  separate. 
Rev.  in  Centralb.  f.  Bakt.,  Bd.  xvi,  1894,  pp. 
965-967. 

('95).  DAVENPORT,  C.  B.,  AND  CASTLE,  W.  E.    On  the 
acclimatization   of  organisms  to  high  tem- 
peratures.      Archiv.     f.     Entwickelungsme- 
chanik  der  Organismen,  Bd.  n,  Heft  2,  1895, 
pp.  227-249.    Bibliography  of  3  pages. 
Organisms  become  acclimatized  to  high  temperature  by 
losing  water  from  their  protoplasm. 


CHANGES    DUE   TO   ENVIRONMENT;    CULTURE-MEDIA. 


223 


('97).  PECKHAM,  ADELAIDE  WARD..  The  influence  of 
environment  upon  the  biological  processes 
of  the  various  members  of  the  colon  group 
of  bacilli.  Jour,  of  Exper.  Medicine,  1897, 
Sept.,  pp.  S49-S9I. 

('97).  AUERBACH,  WILHELM.  Ursachc  der  Hem- 
mung  der  Gelatine-Verfliissigung  duroh 
Bacterien  durch  Zuckerzusatz.  Arch.  f. 
Hygiene,  Bd.  xxxi,  1897,  pp.  311-318. 

('98).  PICKER,  MARTIN.  Ueber  Lebensdauer  und 
Absterben  yon  pathogenen  Keimen.  (Habili- 
tationsschrift.)  Leipzig,  Veit  &  Comp, 
1898.  Rev.  in  Centralb.  f.  Bakt.,  xxvn  Bd., 
1900,  p.  685. 

Sorts  of  glass  used  for  holding  culture  media  have  a 
distinct  influence  on  certain  bacteria  by  reason  of  sub- 
stances dissolved  out  of  it. 

('98).  RuziCKA,  SYANISLAV.  ExperLrnentelle  Studien 
iiber  die  Variabilitat  wichtiger  Charaktere 
des  B.  pyocyaneus  und  des  B.  fluorescens 
liquefaciens.  Centralb.  f.  Bakt,  xxiv  Bd., 

1898,  pp.  11-17. 

('98).  NIEDERKORN,  ERMINIO.  Vergleichende  Unter- 
suchung  iiber  die  versohiedenen  Varietaten 
des  Bacillus  pyocyaneus  und  des  Bacillus 
fluorescens  liquefaciens.  (Inaug.  Diss.). 
Freiburg,  Switzerland,  1898.  Rev.  in  Cen- 
tralb. f.  Bakt.,  xxvii  Bd.,  1900,  pp.  749-750. 

('98).  LONDON,  E.  S.  Le  microbiometre  et  son  appli- 
cation a  1'etude  des  phenomenes  d'inanition 
ohez  les  foacteries.  Arch,  des  sci.  biol.  pub- 
liees  par  1'inst.  imp.  de  med.  exper.  a  St. 
Petersburg,  T.  vi,  1898,  pp.  71-80. 

The  bacteria  experimented  on  endured  starvation  only 
49  to  88  days. 

('99).  HELLSTROEM,  F.  E.  Zur  Kenntnis  der  Einwir- 
kung kleiner  Glukosemengen  auf  die  Vital- 
itat  der  Bakterien.  Centralb.  f.  Bakt.,  xxv 
Bd.,  1899,  pp.  170-180  and  217-223. 

Even  small  quantities  of  grape  sugar  proved  harmful 
iu  case  of  acid-forming  bacteria. 

('99).  MADSEN,  TH.  Einige  Bemerkungen  zu  dem 
Aufsatz  yon  F.  E.  Hellstrom  zur  Kenntnis 
der  Einwirkung  kleiner  Glukosemengen  auf 
die  vitalitat  der  Bakterien.  Centralb.  f. 
Bakt.,  xxv  Bd.,  1899,  pp.  712-713. 

('99).  TOMASCZEWSKI,  EGON.  Ueber  das  Wachstum 
der  Tuberkelbacillen  auf  kartoffelhaltigen 
Nahrboden.  Zeitschr.  f.  Hyg.  Bd.  xxxn, 

1899,  Heft  2,  p.  247. 
('oo).  KOHLBRUGGE.    See  XLVI. 

Coo).  SMITH,  THEOBALD.  Variation  among  path- 
ogenic bacteria.  Jour.  Best.  Soc.  Med.  Sci., 
vol.  iv,  No.  5,  1900,  pp.  95-109. 

('oo).  EMMERICH,  RUDOLF,  AND  SAIDA.  Ueber  die 
morphologischen  Veranderungen  der  Milz- 
brandbacillen  'bei  ihrer  Auflosung  durch 
Pyocyanase.  Centralb.  f.  Bakt.,  xxvn  Bd., 

1900,  pp.  776-787,  with  i  colored  plate, 
('oo).  KRAUSE,    PAUL.     Beitrage   zur   Kenntnis    des 

Bacillus    pyocyaneus.      Centralb.    f.    Bakt., 
xxvn  Bd.,  1900,  pp.  769-775. 

Considers  effect  of  electricity  (Tesla  stream);  pigment 
formation  when  in  symbiosis  with  streptococci  ;  beha- 
vior in  hydrogen,  carbon  dioxide,  illuminating  gas,  hy- 
drogen sulphid  ;  in  vacuo  ;  nature  of  the  pigments. 

Coo).  SMITH,  THEOBALD.  Die  Bedeutung  von  Varie- 
taten bei  pathogenen  Bakterien.  Centralb. 
f.  Bakt.,  xxvii  Bd.,  1900,  pp.  676-677. 


('oo).  MATZUSCHITA,  TEISI.  Ueber  die  Verander- 
lichkeit  der  Eigenschaft  des  Bacillus  an- 
thracis,  Gelatine  zu  verfliissigen.  Cenitralb. 
f.  Bakt.,  xxvni  Bd.,  1900,  pp.  303-304. 

Coo).  MATZUSCHITA,  TEISI.  Die  Einwirkung  des 
Kochsalzgehaltes  des  Nahrbodens  auf  die 
Wuchsform  der  Mikroorganismen.  Zeits.  f. 
Hyg.,  1900,  Bd.  xxxv,  p.  495. 

Coi).  ROSENFELD,  A.  Ueber  die  InvoJutionsformen 
ewu'ger  pestahnlicher  Bakterien  auf  Kooh- 
salzagar.  Centralb.  f.  Bakt.,  I  Abt,  Bd. 
xxx,  1901,  pp.  641-653. 

('02).  SMITH,  THEOBALD.  The  relation  between 
•bovine  and  human  tuberculosis.  The  Medi- 
cal News,  New  York,  vol.  LXXX,  Feb.  22, 
1902,  pp.  342-346.  Also  a  separate,  pp.  14. 

('02).  LEPOUTRE,  L.  Reoherches  sur  la  transforma- 
tion experimentale  de  bacteries  banales  en 
races  parasites  des  plantes.  Annales  de  1'inst. 
Pasteur,  T.  xvi,  1902,  pp.  304-312. 

('04).  MAASSEN,  ALBERT  Die  teratologischen 
Wuchsformen  (Involutionsformen)  der 
Bakterien  und  ihre  Bedeutung  als  diagnos- 
tisches  Hilfsmittel.  Arbeiten  a.  d.  Kaiser- 
lichen  Gesundheitsamte,  Berlin,  1904,  Bd. 
xxi,  Heft  3,  pp.  385-400,  pi.  x  to  xv. 


XVI.    Culture-Media. 

('59).  PAYEN.    Sur  la  gelose  et  les  nids  de  salangane. 
C.  R.  des  se.  de  1'Acad.  des  sci.,  Paris,  T. 
XLIX,  17  Oct.,  1859,  pp.  521-530. 
According  to  Payen.  agar-agar  has  the  following  aver- 
age composition  :  Carbon,  42.770;  hydrogen, 5.771;;  oxveen 
51.445;  total,  100.000. 

('82) .  iNAEGELi,  C.  v.      Untersuchungen  ueber  Niedere 

Pilze.  a.  d.  Pflanzenphys.  Inst.  i.  Muenchen. 

1882,  pp.  1-285. 
('86).  GUILLEBEAU,   A.,   ET   DE   FREUDENREICH,   ED. 

Preparation  des  gelees  a  base   d'agar-agar. 

Archiv.  des  sci.  phys.  et  nat.,  3e  Periode,  T. 

xv,  Geneve  1886,  pp.  466-468. 

This  author  describes  a  method  of  making  agar  with- 
out filtering.  Cook  for  a  quarter  of  an  hour  a  water 
solution  of  agar  (2  per  cent),  salt  0.5  per  cent,  and  pep- 
tone i  per  cent.  This  is  alkaline ;  neutralize.  Add  an 
equal  quantity  of  bouillon  prepared  the  day  before 
This  bouillon  contains  salt  and  peptone  in  the  same  pro- 
portion (i  kilo,  of  meat  for  2  litres  of  water).  Cook  in 
the  autoclave  for  two  hours  at  a  temperatuoe  of  120°  to 
125°  C.  Do  not  pass  130°,  as  agar  changes  color.  Remove 
and  let  stand.  After  4  to  5  hours  one  has  in  the  upper 
part  of  the  flask  a  limpid  liquid  with  all  impurities  at 
the  bottom.  During  this  time  the  temperature  must  be 
kept  above  42°.  Decant  and  sterilize  at  110°  C. 

('86).  MIQUEL,  P.  "De  la  culture  des  bacteries"  in 
Septieme  Memoire  sur  les  organismes  micro- 
scopique  de  1'air  et  dex  eaux.  Annuaire  de 
1'observ.  a  Montsouris  pour  1885.  See  espe- 
cially pp.  569-570. 

This  deals  with  use  of  Fucus  crispus  as  basis  for  a  solid 
culture-medium. 

('87).  SCHOTTELIUS,  M.  Einige  Neuerungen  an  bac- 
teriologischen  Apparaten.  2  Vollstandig 
klarer  Agar-Nahrboden.  Centralb.  f.  Bakt., 
1887,  II  Bd.,  pp.  100-101. 

Agar  is  soaked  about  five  minutes  in  2  per  cent  hydro- 
chloric acid  and  then  washed  in  frequent  changes  of 
water.  Five  to  ten  percent  of  this  agar  is  then  macerated 
over  night  in  bouillon  at  room  temperature ;  cooked  ; 
peptone  and  salt  added  ;  neutralized  with  carbonate  of 
soda  orpotash  ;  again  cooked, and  finally  passed  through 
filter  paper. 


224 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('87).  DAI,  Pozzo,  D.  Das  Eiweiss  der  Kiebitzeier 
als  Nahrboden  fiir  Mikroorganismen.  Med. 
Jahrb.  Jahrg.,  1887,  pp.  523-529. 

('87).  ABBOTT,  A.  C.  An  improvement  in  the  method 
of  preparing  blood  serum  for  use  in  bac- 
teriology. Medical  News,  1887,  vol.  i,  p. 
207.  Rev.  in  Centralb.  f.  Bakt,  n  Bd.,  1887, 
pp.  424-425. 

('87).  RASKIN,  M.  Zur  Zucbtung  .der  pathogenen 
Mdkroorgani'smen  auf  aus  Milch  bereiteten 
festen  und  durchsichtigen  Nahrboden,  St. 
Petersburger  med.  Wochenschr,  xn  Jahrg., 
1887,  pp.  357-360. 

('88).  VON  FREUDENREICH,  E.  Zur  Berettung  des 
Agar-Agar.  Centralb.  f.  Bakt.,  1888,  in 
Bd.,  pp.  797-798. 

Recommends  filtering  nutrient  agar  in  the  autoclave  at 
about  I  io°C.  Time  required  30  to  60  minutes. 

('88).  VAN  PUTEREN.  Ueber  die  Herstellung  von 
festen  Nahrboden  aus  Milch  zu  Mikro- 
organismen  Kulturen.  (Russian.)  1888. 
('88).  HUEPPE,  FERDINAND.  Ueber  die  Verwendung 
von  Eiern  zu  Kulturzwecken.  Centralb.  f. 
Bakt.,  1888,  iv  Bd.,  pp.  80-81. 

Cultures  are  made  in  the  uncooked  eggs  after  shaking 
and  proper  surface  sterilization. 

('88).  Roux,  E.  De  la  culture  sur  pomme  de  terre. 
Ann.  de  1'Inst.  Pasteur,  T.  n,  1888,  pp. 
28-30,  2  figs. 

('89).  PETRI,  R.  J.  Ueber  den  Gehalt  der  Nahrgela- 
itine  an  Salpetersaure.  Centralb.  f.  Bakt.,  v 
Bd.,  1889,  pp.  457-46o. 

Commercial  gelatin  generally  contains  nitrates  in  con- 
siderable quantities. 

('89).  VOIGTLAENDER,  FEIJX.  Ueber  die  Diffusion  in 
Agargallerte.  Zeitschr.  f.  Physik.  Chemie., 
Bd.  in,  1889,  pp.  3i6-335- 

A  study  of  some  of  the  physical  properties  of  agar- 
The  rate  of  diffusion  of  many  acids  and  other  substances 
was  determined.  The  conclusions  are : 

1.  The  diffusion  in  agar  jelly  from  watery  solutions  is 
not  disturbed  by  the  process  of  imbibition. 

2.  The  validity  of  Pick's  law  for  dilute  solutions  was 
amply  demonstrated. 

3.  The  rapidity  c  f  diffusion  of  a  substance  in  different 
concentrations  of  the  affar  ielly  is  the  same.  The  diffusion 
constants  observed  in  the  jelly  are  like  those  of  water  or 
greater  or  less. 

4.  With  rise  of  the  temperature  the  constant  does  not 
increase  in  linear  relation,  but  the  amount  of  the  salt 
which  enters  in  increases. 

('89).  REIN,  J.  J.  The  industries  of  Japan;  together 
with  an  account  of  its  agriculture,  -forestry, 
arts,  and  commerce ;  from  travels  and  re- 
searches undertaken  at  the  cost  of  the  Prus- 
sian Government;  with  44  illustrations  and 
3  maps.  London,  Hudder  &  Stoughton,  27 
Paternoster  Row,  1889,  pp.  xn,  57O.  2d  re- 
vised German  edition,  1905. 

The  cartilaginous  Florideoe,  particularly  species  and 
varieties  of  Gigartinese,  CaulacautheEe,  Gelide,  Sphaero- 
cocceaj,  and  Tylocarpea;,  are  distinguished  for  their  high 
proportion  of  pararabin,  and  furnish,  with  boiling  water 
algee-jelly.  They  are  gathered  in  great  quantity  on  all 
the  coasts  of  the  Malay  Archipelago  and  the  waters  of 
China  and  Japan,  and  are  utilized  in  part  direct  as  food, 
partly  in  the  preparation  of  algse-glue,  Jap.  Fu-nori,  or 


tne    iviaiay   wuiu  /i^;u -.-\i^;ii  ,  i.  c.,  vcjgcttijjn-.      ^>.»    uniu*. 

was  originally  applied  to  Gigartina  (Eucheuma)  isifor- 
mis,  G.  spinosa,  and  G.  tenax,  which  is  collected  near 
Singapore,  for  example,  in  great  masses,  and  shipped  to 
China.  The  Chinese  use  them  not  only  for  food  but  make 
of  them  Hal-Thao,  a  transparent  glue,  with  which  they 
stiffen  *ilk  and  other  stuffs,  and  also  fill  up  the  interstices 
of  coarse  clothes  for  the  manufacture  of  lanterns. 


('89).  PETRI,  R.  J.  Nachtrag  zu  "Ueber  den  Gehalt 
der  Nalirgelatdne  an  Salpetersaure."  Cen- 
tralb. f.  Bakt.,  v  Bd.,  1889,  pp.  679-680. 

('90).  KUEHNE,  W.  Kieselsaure  als  Nahrboden  fiir 
Organismen.  Zeitsohr.  f.  Biol.  Neae  Folge, 
Bd.  ix,  Der  ganzen  Reihe,  Bd.  xxvn,  1890, 
pp.  172-179. 

('90).  TISCHUTKIN,  N.  Eine  vereinfachte  Methode 
der  Bereitung  von  Fleisch-Pepton-Agar. 
Wratsch.,  1890,  No.  8,  pp.  177-178.  Reviewed 
in  Centralb.  f.  Bakt.,  Bd.  ix,  p.  208,  1891. 

The  crude  agar-agar  is  first  exposed  for  15  minutes  in 
acetic-acid  water  (5  glacial  acid,  100  water). 

('91).  SCHULTZ,  N.  K.  Zur  Frage  von  der  Bereitung 
einiger  Nahrsubstrate.  Centralb.  f.  Bakt., 
x  Bd.,  1891,  pp.  52-64. 

('91).  SLESKIN,  P.  Die  Kieselsaure  Gallerte  als 
Nahrsubstrat.  Centralb.  f.  Bakt.,  Bd.  x, 
1891,  pp.  209-213. 

('91).  MARPMANN.  Mittheilungen  aus  der  Praxis. 
i,  Ersatz  fiir  Agar;  2,  Ersatz  fiir  Gelatine. 
Centralb.  f.  Bakt.,  x  Bd.,  1891,  pp.  122-124. 

('92).  PETRI,  R.,  UND  MAASZEN,  ALBERT.  Ueber  die 
Bereitung  von  Nahrbouillon  fiir  bakteriolo- 
gische  Zwecke.  Arbeiten  aus  d.  kaiserl. 
Gesundheitsamte,  Bd.  vm,  No.  2,  1892,  pp. 
3II-3I4. 

('92).  LOEW,  O.  Ueber  einen  Bacillus,  welcher 
Ameisensaure  und  Formaldehyd  assimiliren 
kann.  Centralb.  f.  Bakt.,  xn  Bd.,  1892,  pp. 
462-465. 

('92).  DE  LAGERHEIM,  G.  Macaroni  als  fester  Nahr- 
boden. Centralb.  f.  Bakt.,  xi  Bd.,  1892,  pp. 
147-148- 

Author  states  that  cultures  of  chromogenic  bacteria  on 
macaroni  stand  out  from  the  background  very  distinctly 
and  are  very  instructive.  The  whitest  macaroni  should 
be  selected.  It  is  cut  into  pieces  4.5  cm.  long.  These  are 
put  into  test  tubes  and  covered  i  cm.  over  with  water  and 
cooked  for  15  minutes.  The  water  is  then  carefully  poured 
off  and  the  media  sterilized  in  streaming  steam  in  the 
usual  way. 

('92).  SEILER,  F.  Influence  de  la  composition  de  la 
gelatine  nutritive  sur  le  developpement  des 
colonies  microbiennes.  Schweizerische 
Wochenschr.  f.  Chemie  u.  Pharm.,  1892, 
pp.  261-263. 

('92).  SCHUTZ,  J.  Q.  A  rapid  method  of  making 
nutrient  agar.  Bull.  Johns  Hopkins  Hos- 
pital,vol.  in,  July-Augusit,  1892,  p.  92. 

A  useful  and  easy  method. 

('93).  USCHINSKY.  Ueber  eine  eiweissfreie  Nahr- 
losung  fur  pathogene  Bakterien,  nebst 
einigen  Bemerkungen  fiber  Tetanusgift. 
'Centralb.  f.  Bakt.,  Bd.  xiv,  1893,  No.  10,  pp. 
316-319- 

('93).  NASTIUKOFF.  Ueber  Nahrboden  aus  Eigelb 
fur  Bakterienkulturen.  Wratsch.,  1893,  No. 
33  and  34.  Rev.  in  Centralb.  f.  Bakt.,  xvn 
Bd.,  1895,  pp.  492-493- 

('93).  HESSE,  W.  Ueber  den  Einfluss  der  Alkalescenz 
des  Nahrbodens  auf  das  Wachsthum  der 
Bakterien.  Zeitschr.  f.  Hygiene,  Bd.  xv, 

1893,  pp.  183-191-    3  plates. 

('94).  FRAENKEL,  C.  Beitrage  zur  Kenntniss  des 
Bakterienwachsthums  auf  eiweissfreien 
Nahrlosungen.  Hyg.  Rumlschau,  Jahrg.  iv, 

1894,  pp.  769-776. 


CULTURE-MEDIA. 


225 


('94).  WESCNER,  F.     Die  Bereitung  eines  festen  un- 
•durchsiditigcn     Nahrbodens    fur    Bakterien 
aus    Hiihnerciorn.      Centralb.    f.    allg.    Path, 
u.  path.  Anat.,  Bd.  v,  1894,  pp.  57-59. 
Eggs  are   shaken  until  the  yolks  and  whites  are  thor- 
oughly mixed.     They  are  then  boiled  hard.     The  shell  is 
now  removed,  the  egg  cut  into  suitable  pieces,  put  into 
test  tubes  and  treated  exactly  like  potato  cylinders.    This 
media  is  said  to   give  very  characteristic  growths  with 
many  bacteria. 

('95).  TuRRO,  R.  Ueber  Stroptokokkenziichtung  auf 
sauren  Na'hrboden.  Centralb.  f.  Bakt,  Bd. 
xvir,  i  Abt.,  1895,  pp.  865-874. 

('95).  SEDGWICK  AND  PRESCOTT.    See  XLVI. 

('95).  ELSNER.  Untersuchungen  iibar  eleotives 
Wachsthum  der  Bacterium  coli-Arten  und 
des  Typhusbacillus  und  dessen  diagnostiche 
Verwerthbarkeit.  Zeitschr.  f.  Hyg.,  Bd.  xxi, 
1895,  pp.  25-31. 

("95).  SMITH,  THEOBALD.  Ueber  die  Bedeutung  des 
Zuckers  in  Kulturmedien  fur  Bakterien. 
Centralb.  f.  Bakt.,  xvin  Bd.,  1895,  pp.  1-9. 

('95).  BLEISCH,  MAX.    See  xvn. 

('95).  HAEGLER,  CARL  S.     Zur  Agarbereitung.    Cen- 
tralb.  f.   Bakt.,  xvn  Bd.,   189=:,  pp.  558-560, 
with  2  figs. 
Advises  centrifuging  to  clear  the  agar. 

('95).  DEYCKE,    G.      Die    Benutzung    von    Alkalial- 

buminaten  zur  Herstellung  von  Nahrboden. 

Centralb.   f.   Bakt.,  xvn  Bd.,   1895,  pp.  241- 

24S. 
C9S)-  FULLER,  GEO.  W.    On  the  proper  reaction  of 

nutrient     media     for    bacterial     cultivation. 

Jour.   Am.    Pub.    Health    Asso.,   Oct.,    1895, 

vol  xx. 

A  very  useful  paper.  It  is  recommended  for  general 
reading. 

('95).  HEIM,  L.  Zur  Bereitungsweise  von  Nahr- 
mitteln.  Centralb.  f.  Bakt.,  xvn  Bd.,  1895, 
pp.  190-195,  i  phot. 

('95).  ZUPNIK,  LEO.  Zur  Agarbereitung.  Centralb. 
f.  Bakt.,  xvin  Bd.,  1895,  p.  202.  Review  in 
Bot  Centralb.,  Bd.  LXV,  1896,  p.  52. 

Author  obtains  clear  agar  by  filtering  it  through  a  thin 
layer  of  absorbent  cotton,  placed  funnel-form  in  the  hot 
water  filter,  wet  with  hot  distilled  water  and  pressed  in 
place  with  the  fingers.  The  bouillon  must  be  clear  to 
start  with.  The  flgar  powder  is  then  added  and  cooked 
i  hour  in  streaming  steam. 

('95).  MAASSEN,   ALBERT.     Die  organischen   Sauren 

als     Nahrstoffe     und     ihre     Zersetzbarkeit 

durch    die    Bakterien.     Arb.    a.    d.    kaiserl. 

Gesundheitsamte,    Bd.    xu,    Zweites    Heft, 

1895,  PP-  340-411- 
('96).  CAPALDI,    ACHILLE.      Zur    Verwendung    des 

Eidotters   als    Nahrbodenzusatz.      Cenitralb. 

f.  Bakt.,  xx  Bd.,  1896,  pp.  800-803. 
('97).  HESSE.    See  XLVI. 
('97).  MARPMANN.    See  XLIX. 
('97).  FORSTER,  J.    Niihrgelatine  mit  hohem  Schmelz- 

punkte.     Centralb.   f.  Bakt.,  xxn  Bd.,   1897, 

pp.  341-343. 

By  careful  minimizing  of  heat  the  author  obtains 
sterile  nutrient  gelatin  melting  at  290-30°  C. 

('97).  STODDART,  F.  WALLIS.  New  method  of  sepa- 
rating the  typhoid  bacillus  from  the  bacillus 
coli  communis,  with  notes  on  some  tests  for 
the  typhoid  bacillus  in  pure  cultures.  Uni- 
versity College,  Bristol.  The  Jour,  of  Path- 
ology and  Bacteriology,  iv,  1896-97,  p.  429. 


('97).  BOKORNY,  TH.  Grenze  der  wirksamen  Ver- 
diiniiung  von  Nahrstoffen  bei  Algen  und 
Pilzen.  Bio.  Centralb.  June  15,  1897,  pp. 
417-426. 

Author  states  that  0.002  per  cent  peptone  serves  no 
longer  as  a  nutrient  for  bacteria,  but  o.oio  per  cent  does. 
In  mineral  solutions,  monopotassium  phosphate,  mag- 
nesium sulphate,  and  calcium  nitrate,  a  dilution  to  o.ooi 
per  cent  still  nourishes  algae,  but  not  bacteria  ;  with  0.005 
per  cent  solutions,  bacteria  appeared. 

('97).  LONDON,  E.  S.  Schnelte  und  leichte  Methode 
zur  Bereitung  des  Nahragars.  Centralb.  f. 
Bakt.,  xxi  Bd.,  1897,  pp.  686-687. 

('97).  Hiss,  PHILIP  HANSON.  On  a  method  of  isolat- 
ing and  identifying  bacillus  ityphosus,  based 
on  a  study  of  bacillus  typhosus  and  members 
of  the  colon  group  in  semi-solid  culture 
media.  Jour.  Exp.  Med.,  vol.  u,  1897,  pp. 
677-700. 

('98).  SMITH, ERWIN  F.  Potato  as  a  culture-medium 
with  some  notes  on  a  synthesized  substitute. 
Proc.  Am.  Asso.  Adv.  Sci.,  Vol.  XLVII,  1898, 
p.  411.  Also  a  separate.  Centralb.  f.  Bakt., 

2  Abt.,  Bd.  v,  p.  102. 

('98).  BOKORNY,  TH.  Sources  of  Carbon  for  Bac- 
teria. See  table  by  Bokorny  in  his  Lehrbuch 
der  Pflanzenphysiologie,  pp.  56-59.  Paul 
Parey,  Berlin,  1898. 

('98).  GIESENHAGEN,  K.  Eine  Vorrichtung  zum 
Filtrieren  von  Nahragar.  Centralb.  f.  Bakt., 
xxiv  Bd.,  1898,  pp.  501-502. 

('99).  YOKOTE,  T.  Ueber  die  Darstellung  von 
Nahragar.  Centralb.  f.  Bakt.,  xxv  Bd.,  1899, 
PP-  379-38o. 

Author  heats  his  filtered  bouillon,  to  which  agar  has 
been  added,  for  i  hour  on  a  sand  bath,  after  which  it 
filters  readily  if  the  sand  temperature  has  been  above  110° 
C.  It  will  not  filter  satisfactorily  if  lower  temperatures 
are  used. 

('99).  BLIESENER.  Ueber  Gelatinekulturen  im  Briit- 
schrank.  Zeitsohr.  f.  Hyg.,  Bd.  xxxii,  Heft 
i,  1899.  Rev.  in  Centralb.  f.  Bakt.,  xxvn 
Bd.,  1900,  pp.  472-473- 

Author  prepares  a  gelatin  which  remains  solid  at 
27-30°  C. 

('99).  CESARIS-DEMEL.       Udber     das     verschiedene 
Verbal/ten  einige  Mikroorganismen  in  einem 
gefarbten   Nahr-Mittel.     Centralb.    f.   Bakt., 
xxvi  Bd.,  1899,  pp.  529-540,  with  2  plates. 
The  medium  recommended  is  liver  broth  with  tincture 
of  litmus.     This  serves,  it  is  said,  to  differentiate  certain 
species. 

(.00).  GLAESSNER,  PAUL.     Ueber  die  Verwertbarkeit 
einiger  neuer  Eiweisspraparate  zu   Kultur- 
zwecken.    Centralb.  f.  Bakt.,  XXVH  Bd.,  1900, 
pp.  724-732. 
Comparative  tests  of  somatose,  nutrose,  etc. 

('02).  YENDO,  K.  Uses  of  marine  Algae  in  Japan. 
Postelsia,  Sit.  Paul,  Minn.,  1902,  pp.  3-18. 

3  plates  and  3  Japanese  prints. 

Gelidium  corneum  (Japanese  Ten-gusa)  furnishes  the 
agar-agar  of  commerce. 

('02).  WHIPPLE,  GEORGE  C.  On  the  physical  prop- 
erties of  gelatin,  with  reference  to  its  use 
in  culture  media.  Technology  Quarterly, 
Boston,  Mass.,  vol.  xv,  pp.  127-160.  Also  a 
separate. 

('04).  GAGE,  STEPHEN  DE  M.,  AND  ADAMS,  GEORGE  O. 
Studies  of  media  for  the  quantitative  estima- 
tion of  bacteria  in  water  and  sewage.  Jour, 
of  Infectious  Diseases,  vol.  i,  No.  2,  1904, 
PP-  358-377.  Also  a  separate. 


226 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('04).  HESSE,  GUSTAV.  Beitrage  zur  Herstellung  von 
Nahrboden  und  zur  Bakterienziichitung. 
Ztschr.  f.  Hyg.,  1904,  Bd.  XLVI,  pp.  1-22. 

Discusses  effect  of  adding  various  acids  and  alkalies, 
effect  of  heat  on  reaction,  use  of  insoluble  glass,  etc. 

Most  of  the  methods  advised  are  already  in  use  in  many 
laboratories  in  the  United  States. 


XVII.    Methods  or  Work,  Apparatus,  Etc. 

('73).  KLEBS,  E.    Beitraege  zur  Kenntniss  der  Micro- 
coccen.    Arch.  f.  exp.  Path.  u.  Plharmakol., 
1873,  Bd.  i,  pp.  31-64,  with  4  plates. 
Klebs  here  outlines  his  fractional  method  of  culture. 
See  especially  pp.  46-47. 

('77).  KOCH.    See  LV. 

('78).  LISTER,  JOSEPH.     On  the   lactic   fermentation 

and  its  bearings  on  pathology.    Trans.  Path. 

Soc.  of  London,  1878,  vol.  xxix,  pp.  425-467. 

Lister's  dilution  method  for  obtaining  pure  cultures  is 

here  described.    See  especially  page  445  et  seq. 

('81).  KOCH,  ROBERT.  Zur  Untersuchung  von  path- 
ogenen  Organismen.  Mittheil.  a.  d.  kaiser- 
lichen  Gesundheitsamte,  Bd.  I,  1881.  Ber- 
lin, pp.  1-48,  14  plates. 

In  this  paper  Koch  outlined  his  now  universally  used 
method  of   obtaining  pure   cultures  (colonies)  on  solid 


was  assumed  that  the  quantity  used  to  inoculate  each 
flask  contained  only  one  organism  (p.  19). 

('83).  KOCH,  ROBERT.     Ueber  die  Milzbrandimpfung, 
eine   Entgegnung   auf  den   von   Pasteur   in 
Genf  gehaltenen  Vortrag.     Kassel  und  Ber- 
lin, Theodor   Fischer,    1882.     The   same   in 
French.    Theo.  Fischer,  1883. 
The  celebrated  Koch  "  rules  of  procedure  "  are  given  in 
this  paper. 

('84).  HOFFMANN,    F.    W.     Einfacher    Einbettungs- 

apparat.     Zoologisoher  Anzeiger,  vii  Jahrg., 

1884,  No.  157-184,  Leipzig,  pp.  230-232,  I  fig. 

Makes  use  of  a  vacuum  to  hasten  the  infiltration  of 

paraffin. 

('84).  ERRERA.  Sur  1'emploi  de  1'encre  de  phine  en 
Microscopic.  Bull.  Soc.  Beige  d.  Microscop., 
1884. 

Not  seen.    The  ink  is  used  to  form  a  contrasting  back- 
ground. 

('86).  VON  ESMARCH,  ERWiN.  Ueber  eine  Modifika- 
ition  des  Koch'sohen  Plattenverfahrens  zur 
Isolierung  und  zum  quantitativen  Nachweis 
von  Mikroorganismen. — Zeiitsoh.  f.  Hygiene, 
1886,  Bd.  i,  pp.  293-301,  3  figs. 
The  method  of  roll  cultures  is  here  described. 

('87).  PETRI,  R.  J.  Eine  kleine  Modification  <les 
Kooh'schen  Plattenverfahrens.  Centralb.  f. 
Balct,  i  Bd.,  1887,  No.  9,  pp.  279-280. 

Description  of  the   now  everywhere   common    Petri- 
dishes. 

('87).  ESMARCH.  See  xxxin. 

('89).  BEYERINCK,  M.  W.  L'auxanographie  ou  la 

methode  de  -1'hydrodiffusion  dans  la  gelatine 

appliquee  aux   recherches  mdcrobiologiques. 

Arch,  neertandaises  de  sci.  ex.  et  nat.,  1889, 

T.  xxni,  pp.  367-372. 
('90).  KUEBLER.  Untersuchungen  iiber  die  Brauch- 

barkeit  der  "Filtres  sans  pression,  Systeme 

Chamberland-Pasteur."     Zeitschr.    f.    Hyg., 

Bd.  VHI,  1890,  pp.  48-54. 


('91).  NUTTALL,  GEORGE  H.  F.  A  method  for  the 
estimation  of  the  actual  number  of  Tubercle 
bacilli  in  tuberculous  sputum.  With  a  note 
on  the  general  application  of  the  method  to 
bacteriology.  Bull,  of  the  Johns  Hopkins 
Hospital,  vol.  n,  1891,  pp.  67-76. 

By  use  of  this  method  flasks  or  tubes  of  media  can  be 
inoculated,  it  is  said,  with  predetermined  exact  num- 
bers of  bacteria. 

('91).  BUJWID,  O.  Eine  einfaohe  Filtervorrichtung 
zum  Filtriren  sterilisirter  Fliissigkeit.  Cen- 
tralb. f.  Bakt.,  ix  Bd.,  1891,  pp.  4-5,  mit  i 
Abbildung. 

('91).  GABRITSCHEWSKY,  G.  Zur  Technik  der  foak- 
teriologischen  Untersuchungen,  I.  Graduirte 
Kapillarpipetten  zum  Abmessen  sehr  kleiner 
Fliissigkeitsmengen.  Centralb.  f.  Bakt.,  x 
Bd.,  1891,  pp.  248-250,  mit  2  figs. 

('91).  NORDTMEYER,  H.  Ueber  Wassernltration  durch 
Filter  aus  gebranniter  Infusorienerde. — 
Zeitsch.  f.  Hygiene,  1891,  Bd.  x,  pp.  145- 
154,  i  fig- 

('91).  SMITH,  THEOBALD.  Kleine  bakteriologische 
Mittheilungen.  Centralb.  f.  Bakt.,  x  Bd., 

1891,  pp.  177-186,  mit  2  figs. 

First  two  notes  relate  to  apparatus;  the  second,  to  his 
inside-out  method  of  filtering  through  Chamberland 
bougies  (figured). 

('91).  KNAUER,  FRIEDRICH.  Eine  bewahrte  Methode 
zur  Reinigung  gebrauchter  Objekttrager 
und  Deckglaschen.  Centralb.  f.  Bakt.,  Bd. 
x,  1891,  pp.  8-9. 

Even  old  stuck-together  slides  are  said  to  be  cleaned 
perfectly  by  boiling  30  miuutes  in  10  per  cent  lysol,  with 
occasional  stirring.  They  should  then  be  flooded  with 
tap  water,  and  finally  wiped  with  a  soft,  clean,  oil-free 
cloth.  "  In  recent  preparations,  14  days  old,  I  obtained 
even  by  15  minutes  boiling  in  5  per  cent  lysol  solution  a 
perfect  cleaning  of  the  glass." 

('91).  ALTMANN,  P.  Thermoregulator  neuer  Kon- 
struktion.  Cenitralb.  f.  Bakt.,  ix  Bd.,  1891, 
pp.  791-792. 

('91).  BEYERINCK,  M.  W.  Die  Kapillarhebermikro- 
skopirtropfenflasche.  Centralb.  f.  Bakt.,  ix 
Bd.,  1891,  pp.  589-590,  mit  i  Abbildung. 

An  easily  constructed  useful  water  flask,  furnishing 
drops  of  any  size  (see  fig.  16)  or  a  fine  stream. 

('92).  HASWELL,  WM.  A.     On  a  simple  method  of 
substituting    strong   alcohol    for    a    watery 
solution    in   the   preparation   of   specimens. 
Proc.  Linn.  Soc.  N.  S.  W.,  vol.  vi,  for  1891, 
PP.  433-436,  i  fig.    Sydney,  1892. 
This  method  was  worked  out  10  years  later,  indepen- 
dently, in  my  laboratory,  by  Deane  B.  Swingle,  who  had 
his  paper  and  drawings  ready  for  publication  when  this 
earlier  paper  was  discovered.    Swingle's  device,  which  is 
a  considerable  improvement  ou  Haswell's,  is  shown  ill 
Fig.  146.     By  means  of  the  expanded  base  of  the  inner 
tube  a  quite  equable  diffusion  of  the  alcohol  is  secured. 

('92).  GILT  AY,  E.,  u.  ABERSON,  J.  H.  Methode  zur 
Priifung  von  Filtereinrichtungen  wie  die 
Chamberland-Bougies.  Cenitralb.  f.  Bakt., 

1892,  Bd.  xii,  pp.  92-9S,  i  fig- 

('92).  v.  FREUDENREICH,  ED.  Ueber  die  Durchlas- 
sigkeit  der  Chamberland'schen  Filter  fur 
Bakterien.  Centralb.  f.  Bakt.,  xn  Bd.,  1892, 
pp.  240-247. 

The  author's  experiments  lead  to  the  conclusion  that 
the  Chamberlaud  filter  cannot  be  used  safely  more  than  a 
week  for  the  continuous  filtration  of  water.  It  must 
then  be  sterilized. 


METHODS   OF   WORK,    APPARATUS,    ETC. 


227 


('92).  SMITH,  THEOBALD,  AND  MOORE,  V.   A.     Zur 

Pruning    der    Pasteur-Chamberland    Filter. 

Centralb.  f.  Bakt.,  xn  Bd.,  1892,  pp.  628-629, 

mil  i  Abbildung. 
('92).  ALTMANN,    P.      Ein    neuer    Thermoregulator 

fiir     Petroleumheizung    bci     Thermostaten. 

Central),  f.  Bakt.,  xn  Bd.,  1892,  pp.  654-655, 

mit  2  figs. 

('92).  ARLOING,  G.  De  1'influence  des  filtres  mineraux 
sur  les  liquides  contenant  des  substances 
d'origine  microbienne.  C.  R.  des  se.  de 
1'Acad.  des  sea.,  T.  cxiv,  1892,  pp.  1,455-1,457. 

('92).  DE  FREUDENREICH.  De  la  permeabilite  des 
filtres  Chamberland  a  1'egard  des  bacteries. 
Annales  de  Micrographie,  1892,  Tome  iv, 
PP-  SS9-568. 

('92).  HOLM,  JUST  CHR.  Sur  les  methodes  de  culture 
pure  et  specialment  sur  la  culture  sur  plaques 
de  M.  Koch  et  la  limite  des  erreurs  de  cette 
methode.  Meddelelser  fra  Carlsberg  Lab. 
Tredle  Bind.  Andet  Hefte.  Kjobenhavn, 
1892,  pp.  1-23,  Danish  edition,  Tredie  Bind, 
Forste  Hefte,  1891,  pp.  1-32. 

('92).  NENCKI  AND  ZAWADZKI.    See  XLVII. 

('92).  ALTMANN,  P.  Neue  Mikrogaslampen  als 
Sicherheitsbrenner.  Centralb.  f.  Bakt.,  XH 
Bd.,  1892,  pp.  786-787,  3  figs. 

('93).  ZETTNOW.  Reinigung  von  neuen  Deckglasern. 
Centralb.  f.  Bakt.,  Bd.  xiv,  1893,  pp.  63-64. 

The  last  traces  of  fat  are  best  removed  from  covers  by 
burning.  The  clean  covers  are  placed  on  an  8  to  10  cm. 
square  piece  of  sheet  iron,  which  is  then  heated  for  a  few 
minutes  in  the  open  flame  of  a  Bunseu  burner. 

('93).  SCHEPILEWSKY,  E.  A.  Ein  Regulator  zum 
Thermostaten  mit  Wasserheizung.  Cen- 
tralb. f.  Bakt.,  Bd.  xiv,  1893,  pp.  131-138, 
mit  i  fig. 

('93).  GRUBER,  MAX.  Gesichtspunkte  fiir  die  Priifung 
und  Beurteilung  von  Wasserfiltern.  Cen- 
tralb. f.  Bakt.,  Bd.  xiv,  1893.  pp.  488-493. 

('93).  KIRCHNER,  MARTIN.  Gesiohtspunkte  fiir  die 
Priifung  und  Beurteihing  von  Wasserfiltern. 
(Entgegnung  auf  die  gleichnamige  Arbeit 
von  Pro!  Max  Gruber  in  Wien).  Centralb. 
f.  Bakt.,  Bd.  xiv,  1893,  pp.  516-527. 

('93).  BRUNNER,  G.,  AND  ZAWADZKI,  A.  Zahlplatte 
zu  den  Petri'schen  Schalen.  Centralb.  f. 
Bakt.,  Bd.  xiv,  1893,  pp.  616-618,  mit  i  fig. 

A  method  is  given  for  dividing:  a  circle  locm.  in  diame- 
ter into  64  equal  parts.  The  circles  and  sections  are  to  be 
in  white  ink  on  black  paper. 

('93).  SCHOEFER,  HANS.  Ueber  das  Verhalten  von 
pathogenen  Keimen  in  Kleinfiltern.  Cen- 
tralb. f.  Bakt.,  Bd.  xiv,  1893,  pp.  685-693. 

('93).  LAFAR,  FRANZ.  Eine  neue  Zahlvorriohtung 
fur  Plattenkulturen  in  Petrischalen.  Zeitsoh. 
f.  Nalmmgsmitteluntersuchung  u.  s.  w., 
Wien,  1893,  No.  24,  p.  429.  Rev.  in  Centralb. 
f.  Bakt.,  Bd.  xv,  1894,  pp.  331-333,  mit  i  fig. 

('94).  LOEFFLER,  F.  Eine  sterilisirbare  Injections- 
spritze.  Centralb.  f.  Bakt.,  Bd.  xvi,  1894, 
pp.  729-73L 

To  be  had  from  Wittig,  instrument  maker  in  Greifs- 
wald,  Germany. 

('94).  HOUSTON,  A.  C.  Note  on  a  simple  apparatus 
for  collecting  samples  of  water  for  bac- 
teriological purposes,  at  different  depths 
from  the  surface.  Jour,  of  Path,  and  Bact., 
vol.  II,  1894,  pp.  496-497,  i  fig. 


('94).  VALLIN,  E.  Le  regeneration  par  agents 
chimiques  des  filtres  Chamberland.  Revue 
d'hygiene  eit  de  police  sanitaire,  1894,  No. 
11,  p.  946.  Rev.  in  Centralb.  f.  Bakt.,  xvn 
Bd.,  1895,  pp.  496-497. 

Author  recommends  a  sodium  bisulphite  solution 
(1:20)  for  cleaning  niters. 

('94).  KUPRIANOW,  J.    Zur  Methodik  der  keimfreien 
Gewinnung    des    Blutserums.      Centralb.    f. 
Bakt.,  xv  Bd.,  1894,  pp.  458-462,  i  fig. 
Describes  and  figures  a  device  for  filling  test  tubes  with 
exact  amounts  of  fluid  culture  media. 

('94).  FUNCK,  ERNST.  Zur  Frage  der  Reinigung  der 
Deckglaser.  Centralb.  f.  Bakt.,  Bd.  xvi, 
1894,  pp.  113-114. 

Author  cleans  slides  and  covers  which  have  been  used 
with  oil,  Canada  balsam, stains,  etc.,  by  first  placing  them 
for  a  time  in  turpentine,  separating  the  covers  from  the 
slides  as  far  as  possible.  They  are  then  put  into  a  broad 
beaker,  covered  with  hydrochloric  acid,  to  which  is  added 
a  few  grams  (2  to  3  knife-points)  of  chlorate  of  potash. 
This  is  then  covered  with  a  glass  plate  and  heated  on  a 
water  bath  for  a  few  minutes.  The  glasses  are  then 
washed  in  hot  water.  Equal  parts  of  soda,  talcum,  and 
sieved  sawdust  are  now  added  with  water  enough  to 
make  a  thick  fluid  mixture,  which  is  now  heated  for  one- 
half  hour  on  the  water  bath,  with  frequent  shakingof  the 
contents  of  the  beaker.  The  glasses  are  then  washed 
again  in  hot  water,  to  which  a  little  hydrochloric  acid 
has  been  added,  especially,  in  case  some  calcium  carbon- 
ate has  been  deposited  on  the  glass.  Finally  wash  in 
hot  water  or  ethyl  alcohol  and  dry  with  a  soft  cloth. 

('94).  VAN  HEST,  J.  J.  Bakterienluftfilter  und  Bak- 
terienJuftfilterversohluss.  CentraLb.  f.  Bakt., 
Bd.  xvi  1894,  pp.  435-447,  also  pp.  495-499, 
mit  11  figs. 

('95).  KNAUSS,  K.  Eine  einfache  Vorrichtung  zum 
Abfiillen  von  je  10  ccm.  Nahrsubstanz. 
CentraJb.  f.  Bakt.,  xvn  Bd.,  1895,  PP-  878- 
879,  with  i  fig. 

('95).  BANTI,  G.  Eine  einfache  Methode  die  Bak- 
terien  auf  dem  Agar  und  dem  Blutserum  zu 
isolieren.  Centralb.  f.  Bakt.,  xvn  Bd.,  1895 
PP-  556-557. 

Agar  is  slanted,  in  broad  tubes  (diam.  2-3  cm)  The 
material  containing  the  bacteria  is  diluted  to  the  proper 
amount  with  sterile  bouillon  or  water.  The  condensation 
water  is  then  inoculated  by  needle  or  loop,  and  subse- 
quently tilted  over  the  whole  surface  of  the  agar  and 
allowed  to  drain  back  again. 

('95).  BLEISCH,  MAX.  Ein  Apparat  zur  Gewinnung 
klaren  Agars  ohne  Filtration.  Centralb  f 
Bakt.,  xvn  Bd.,  1895,  pp.  360-362. 

(95).  LODE,  ALOIS.  Eine  automatische  Aibfullburette 
fiir  Nahrlosungen  und  Heilserum.  Centralb 
f.  Bakt.,  xvm  Bd.,  1895,  pp.  53-54,  with  3  figs. 

(95).  BRUNNER,  CONRAD.  Notiz  zur  Methode  der 
Isolierung  von  Bakterien  auf  Agarplatten 
im  Reagensglase.  Centralb.  f.  Bakt.,  xvm 
Bd.,  1895,  p.  59- 

(95).  BUJWID,  O.  Bemerkungen  iiber  die  Filtration 
bakterienhailtiger  Flussigkeiten.  Centralb. 
f.  Bakt.,  xvm  Bd.,  1895,  pp.  332-333. 

Considers  the  Chamberland  a  less  fragile  and  safer  filter 
than  the  Berkefeld. 

('95).  ABEL,  RUDOLF.  Ein  Halter  fiir  Objekttrager 
und  Deckglaschen.  Centralb.  f.  Bakt.,  xvm 
Bd.,  pp.  782-783,  with  i  fig. 

('96).  MELNIKOW-RASWEDENKOW,  M.  Ueber  die 
Einstellung  des  d'Arsonvalschen  Thermos- 
taten. Centralb.  f.  Bakt.,  xix  Bd.,  1896,  pp. 
709-712,  with  i  fig. 

('96).  CZAPLEWSKI.    Bakteriologische  Notizen.    Cen- 
tralb. f.  Bakt.,  Bd.  xx,  1896,  pp.  307-313. 
Sixteen  notes  on  methods  of  work. 


228 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('96).  KRETZ,  RICHARD.  Eine  faandliohe  und  leicht 
sterilisierbare  AbfuMvorrichtung  fiir  Kultur- 
flussigkeiten.  Centralb.  f.  Bate.,  xix  Bd., 

1896,  pp.  73-74,  with  I  fig. 

('97).  VAN'T  HOFF,  H.  J.    Eine  schnellere  und  quan- 

titativ  bessere   Methode    der   bakteriologis- 

chen    Plattenzahlung.      Centralb.    f.    Bakt., 

xxi  Bd.,  1897,  pp.  731-733,  with  i  fig. 

Pours  a  suitable  dilution  of  the  bacteria  on  surface 

of  gelatin  or  agar,  and  spreads  fluid  as  widely  as  possible. 

('97).  BOLLEY,  H.  L.  An  apparatus  for  the  bac- 
teriological sampling  of  well  waters.  Cen- 
tralb. f.  Bakt.,  xxn  Bd.,  1897,  pp.  288-290, 
with  I  fig. 

('97).  KASPAREK,  THEODOR.  Bin  Vacuum- Apparat 
zum  Abdampfen  von  Kulturen  mit  Ehmann'- 
scher  Wasserheizung.  Centralb.  f.  Bakt., 
xxn  Bd.,  1897,  pp.  6-7,  with  I  fig. 

('97).  KISCHENSKY,  D.  Ein  Verfahren  zur  schnellen 
mikroskopischen  Untersuchung  auf  Bak- 
terien  in  Deckglas-  und  Objekttrager- 
Praparatem.  Centralb.  f.  Bakt.,  xxi  Bd., 

1897,  pp.  876-877. 

('97).  SMITH,  THEOBALD.  Uetoer  Fehlerquellen  bei 
Prufung  der  Gas-  und  Saurebildung  bei 
Bakterien  und  deren  Vermeidung.  Centralb. 
•f.  Bakt.,  xxii  Bd.,  1897,  pp.  45-49- 

('97).  NOVY,  F.  G.  Neue  Apparate  zum  Filtrieren 
und  zum  Sterilisieren  duroh  Dampf.  Cen- 
tralb. f.  Bakt.,  xxii  Bd.,  1897,  pp.  337-34°, 
with  3  figs. 

('97).  SCHUERMAYER,  B.  Eine  Abanderung  des 
automabischen  Gasabschlusses  beim  Ver- 
loschen  der  Flammen  an  Briitschranken. 
Centralb.  f.  Bakt.,  xxi  Bd.,  1897,  pp.  400- 
401,  with  i  fig. 

('97).  ROBERTSON,  SIGISMUND.  Ueber  Objekttrager- 
und  Deckglashalter.  Centralb.  f.  Bakt.,  xxi 
Bd.,  1897,  pp.  589-591,  with  2  figs. 

('98).  PIORKOWSKI.  Ein  neuer  heizbarer  Parbetisch. 
Deutsch.  med.  Wochenschr.,  1898.  No.  20. 
Rev.  in  Cenitra/lb.  f.  Bakt.,  xxrv  Bd.,  1898, 
pp.  902-903,  i  fig. 

('98).  SMITH,  THEOBALD.  One  of  the  conditions 
under  which  discontinuous  .sterilization  may 
be  ineffective.  Journal  of  Experimental  Medi- 
cine, vol  HI,  1898.  Rev.  in  Centralb.  f.  Bakt., 
xxvi  Bd.,  1899,  p.  585. 

('98).  NOVY,  F.  G.  Ein  neuer  Thermoregulator. 
Centralb.  f.  Bakt.,  xxni  Bd.,  1898,  pp.  1,054- 
1,056,  with  2  figs. 

Made  by  Greiner  andFriedrichs,  Stuetzerbach.Thurin- 
gia,  Germany. 

('98).  MURRILL,  PAUL.  Ein  wirksamer  Gasdruckreg- 
ulator.  Centralb.  f.  Bakt.,  xxni  Bd.,  1898, 
pp.  1,056-1,059,  with  2  figs. 

('99).  KERN,  FERDINAND.  Eine  automattische  Mess- 
pipette  fiir  keimfreie  Flussigkeiten.  Cen- 
tralb. f.  Bakt.,  xxv  Bd.,  1899,  pp.  75-77,  with 
i  fig. 

('99).  NOVY,  F.  G.  Collodium  Sacs.  See  his  book 
entitled  Laboratory  work  in  bacteriology, 
pp.  496-501. 

('oo).  BULLOCH,  WILLIAM.  A  simple  apparatus  for 
obtaining  plate  cultures  or  surface  growths 
of  obligate  anaerobes.  Centralb.  f.  Bakt., 
xxvii  Bd.,  1900,  pp.  140-142,  with  i  fig. 

Author  uses  the  unguentum  resinoe  of  the  British  Phar- 
macopoeia as  a  Ititeing  material.  This  consists  of  resin 
in  powder  200  grams,  or  8  ounces ;  yellow  beeswax  200 
grams,  or  8  ounces  :  olive  oil  200  grams,  or  8  ounces,  and 
lard  iso  grams,  or  6  ounces.  Melt  together  with  gentle 
heat  the  resin  and  wax,  add  the  other  ingredients,  strain 
through  muslin,  and  cool  with  stirring. 


('oo).  WRIGHT,  JAMES  H.  A  simple  method  for 
anaerobic  cultivation  in  fluid  media.  Cen- 
tralb. f.  Bakt.,  xxvn  Bd.,  1900,  pp.  74-75, 
with  i  fig. 

('oo).  STEWART,  C.  BALFOUR.    Apparatus  for  heating 
cultures    to    separate    spore-bearing    micro- 
organisms.    Centralb.    f.   Bakt.,   xxvn   Bd., 
1900,  pp.  366-  367,  with  i  fig. 
This  is  a  modification  of  Meyer's  hot-air  bath. 

('oo).  PETRI,  R.  J.  Eine  einfaohe  Vorrichtung  zum 
Abfiillen  der  Nahrgelatine.  Centralb.  f. 
Bakt.,  xxvii,  Bd.,  1900,  pp.  525-526,  i  fig. 

Coo).  NUTTALL,  GEORGE  H.  F.  Ein  Apparat  zur 
Herstellung  von  Rollkulturen.  Centralb.  f. 
Bakt.,  xxvn  Bd.,  1900,  pp.  605-609,  with  2 
figs. 

('oo).  PIORKOWSKI.  Ein  Apparat  zur  Ermittelung 
von  Desin-fektionswirkungen.  Centralb.  f. 
Bakt.,  xxvii  Bd.,  1900,  pp.  609-610,  with  i  fig. 

Coo).  EPSTEIN,  STANISLAUS.  Ein  neuer  Thermoreg- 
ulator. Centralb.  f.  Bakt.,  xxvm  Bd.,  1900, 
PP-  503-504,  with  I  fig. 

('oo).  PETRI,  R.  J.  Ein  neuer  Reagenzglasstander 
fiir  Kulturen.  Centralb.  f.  Bakt.,  xxvm  Bd., 
1900,  pp.  747-748,  with  i  fig. 

('oo).  SMITH,  R.  GEEIG.  The  measurement  of  bac- 
teria. Proceedings  Linnean  Soc.,  New  South 
Wales  for  1000,  Sydney,  1901,  vol.  xxv,  pp. 
533-536.  Three  figures  in  text.  Also  a 
separate  (issued  Nov.  22,  1900). 

('oo).  PETRI,  R.  J.  Neue  verbesserte  Gelatine- 
Sohalchen  (verbesserte  Petri-Schalchen). 
Centralt).  f.  Bakt.,  xxvm  Bd.,  1900,  pp.  79- 
82,with  3  text  figs. 

This  Petri-dish  has  a  cover  of  yellow  brown  glass,  of 
such  a  form  that  when  they  are  piled  one  above  the  other 
all  are  protected  from  the  action  of  the  violet  rays  of  the 
spectrum.  These  are  made  in  two  forms  by  Paul  Alt- 
111:11111,  Berlin. 

('oo).  PETRI,  R.  J.  Neue  anaerobe  Gelatine-Schal- 
chen-Kultur  (verbesserte  Petri-Schalchen). 
Centralb.  f.  Bakt.,  xxvm  Bd.,  1900,  pp.  196- 
199,  with  2  text  figs. 

('oo).  WRIGHT,  J.  H.  A  simple  method  of  cultivat- 
ing anaerobic  bacteria.  Jour.  Boston  Soc. 
Med.  Sci.,  vol.  v,  1900,  pp.  114-115. 

The  plug  is  pushed  part  way  down  the  test-tube  con- 
taining the  culture.  The  cotton  is  then  partly  saturated 
with  strong  pyrogallic  acid  water  (equal  bulks  of  water 
and  acid).  Sodium  hydrate  solution  (i  NaOH,  2  water)  is 
then  pipetted  on,  and  the  tube  immediately  plugged  air- 
tight with  a  soft  rubber  stopper.  The  media  should  con- 
tain I  percent  glucose.  It  should  be  fresh,  and  its  reac- 
tion not  more  acid  than  +  15.  It  may  be  used  for  fluids, 
or  roll  cultures,  and  other  forms. 

('02).  HILL,  HIBBERT  WINSLOW.  "Hanging  block" 
preparations  for  the  microscopic  observa- 
tion of  developing  bacteria.  Journal  of 
Med.  Research,  Boston,  vol.  vn,  1902  (new 
sen,  vol.  n),  pp.  202-212,  3  figs.  Also  a 
separate. 

('02).  WHERRY,  WM.  B.  Experiments  on  the  per- 
meability of  t*he  Berkefeld  filter  and  the  Pas- 
teur-Chamberland  bougie  to  bacteria  of 
small  size.  Jour,  of  Med.  Research,  vol. 
vni  (n.  s.,  vol.  in),  1902,  pp.  322-328,  I  fig. 

('02).  KELLERMAN,  KARL.  A  method  for  fixing  and 
sectioning  bacteria)  colonies,  fungous  my- 
celium, etc.  Jour.  App.  Micro.,  vol.  v,  1902, 
p.  1,980.  Also  a  separate. 

('02).  KELLERMAN,  KARL  F.  An  improved  method 
for  making  collodion  tubes  for  dialyzing. 
Jour.  App.  Micro.,  vol.  v,  1902,  p,  2,038. 


METHODS   OF   WORK;    MEANS   OF   DIFFERENTIATING. 


229 


fo2).  CARNOT,  PAUL,  ET  GARNIER,  MARCEL.  Sur  la 
technique  des  cultures  en  tubes  de  sable. 
Paris,  C.  R.  soc.  biol,  T.  LIV,  1902,  pp  748- 
750. 

('02).  CARNOT,  PAUL,  ET  GARNIER,  MARCEL.  De 
remploi  des  tubes  de  sable  comme  methode 
generale  d'etude,  d'isolement  et  de  selection 
des  microorganismes  mobiles.  Paris,  C.  R. 
soc.  biol,  T.  LIV,  1902,  pp.  860-863. 

('02).  REGAUD,  CL.  Nouveau  bain  de  paraffine  a 
chauffage  et  regulation  electriques.  J.  anat. 
physiol,  Paris,  T.  xxxvm,  1902,  pp.  193- 
214,  av.  fig. 

('02).  GRIJNS,  G.  Eine  einfache  Vorrichtung,  um 
zu  verhindern,  dass  beim  Gebrauch  des 
Briitapparates  fur  konstante  niedrige  Tem- 
peratur,  System  Lautenschlager  (Katalog 
No.  60,  No.  117),  wenn  das  Eis  im  Behalter 
ausgeht,  das  ungekiihlte  Wasser  in  den 
kalten  Sohrank  fliesst.  Centralb.  f.  Bakt, 
Abt.  i,  Bd.  xxxi,  Originale,  1902,  pp.  430-432. 

('02).  HARRIS,    NORMAN    MACLEOD.    Concerning   an 
improved  method  of  making  collodium  sacs. 
Centralb.  f.  Bakt,  Abt.  I,  Bd.  xxxn,  Orig- 
inale, 1902,  pp.  74-80. 
Dr.  Harris  makes  his  sacs  around  gelatin  capsules. 

(03).  GORSLINE,  CHARLES  S.  On  the  preparation 
and  use  of  collodium  sacs.  Vaughan  Quarter 
Century  Book,  1903,  pp.  390-394.  Ann  Arbor, 
Mich,  George  Walir. 

('03).  WINSLOW,  C.  E.  A,  AND  NIBECKER,  C.  P.  The 
significance  of  bacteriological  .methods  in 
sanitary  water  analysis.  Technology  Quar- 
terly, vol.  xvi,  1903,  pp.  227-239.  Also  a 
separate. 

('04).  REMLINCER,  P.     Le  passage  du  virus  rabique 

a  travers  les  filtres.    2e  mem.  Ann.  de  1'Inst. 

Pasteur,  T.  xvm,  1904,  pp.  150-164. 
("04).  Report  of  the  [English]   committee  appointed 

to  consider  the  standardisation  of  methods 

for  the  bacterioscopic  examination  of  water. 

Journal    of    State    Medicine,    August,    1904. 

See  also   Chemical   News,  vol.   xc,   Oct.   7, 

1904,  pp.  177-179. 

Distilled-water  agar  and  distilled-water  gelatin  are 
recommended  for  use  along  with  nutrient  agar  and 
gelatin.  In  the  search  for  B.  coli  the  committee  recom- 
mends either  the  glucose-formate  broth  of  Pakes  or  the 
bile-salt  broth  of  MacConkey. 


XVIII.    Special  Means  of   Differentiating   Bacteria. 
('84).  GRAM.    See  xiv. 

('87).  VON  ROZSAHEGYI,  A.     Ueber  das  Zuchten  von 
Bakterien  in  gefarhter  Nahrgelatine.     Cen- 
tralb. f.  Bakt.,  ii  Bd.,  1887,  No.  14,  pp.  418-424. 
Author  added  various  substances  to  his  culture  media 
e.  g.  fuchsin,  methylen  blue,  gentian  violet,  vesuvin,  etc. 
Different  bacteria  behave  very    differently    as    respects 
growth,  absorption  of  pigment,  and  change  of  color  in 
the  pigment.     Organisms  may  l>e  differentiated   in   this 
way. 

('88).  BUJVVID,  O.  Neue  Methode  znm  Diagnos- 
ticiren  nnd  Isoliren  der  Choierabakterien 
Centralb.  f.  Bakt.,  1888,  iv  Bd.,  pp.  494-496. 

11124  hours,  in  2  per  cent  feebly  alkaline  peptone  solu- 
tion at  37°  C.,  the  cholera  organism  gives  a  fine  purple-red 
color  on  adding  HC1.  On  longer  cultivation  other  organ- 
isms give  the  same  reaction.  This  depends  on  the  forma- 
tion of  indol  and  a  trace  of  nitrite. 


('90).  PETRUSCHKY,  JOHANNES.  Die  Farbenreaktion 
bakterieller  Stoffwechselprodukte  auf  Lack- 
mus  als  Beitrag  zur  Charakteristik  und  als 
Mittel  zur  Unterscheidung  von  Bakterien- 
arten.  Centralb.  f.  Bakt.,  vn  Bd.,  1890,  pp. 
1-8  and  49-53. 

('92).  Bujwip,  ODO.    Eine  neue  biologische  Reaktion 
fur  die  Choierabakterien.    CentraJb.  f  Bakt 
xn  Bd,  1892,  pp.  59S-S96. 

('92).  BEYERINCK,  M.  W.     Notiz  fiber  die  Cholera- 
rothreaktion.     Centralb.    f.    Bakt,   xn    Bd 
1892,  pp.  715-718. 

('93).  SCHILD.  Formalin  zur  Diagnose  des  Typhus- 
bacillus.  Centralb.  f.  Bakt,  Bd.  xiv,  1893, 
pp.  717-718. 

The  typhoid  organism  will  not  grow  in  bouillon  con- 
taining as  little  formalin  as  1:15,000.  B.  coli  develops 
vigorously  with  as  much  formalin  as  1:3000. 

('93).  MATHEWS,  ALBERT  P.  On  Wurtz's  Method 
for  the  Differentiation  of  Bacillus  typhi 
abdominalis  from  Bacillus  coli  communis, 
and  its  application  to  the  examination  of 
contaminated  drinking  water.  Technology 
Quarterly,  vol.  vi,  1893,  pp.  241-251. 

Litmus  lactose  gelatin  or  agar  is  reddened  by  B.  coli 
and  is  unchanged  or  made  deeper  blue  by  B.  typhosus. 


('93).  GORINI,  KONSTANTIN.  Anmerkung  iiber  die 
Cholerarotreaktion.  Centralb.  f.  Bakt,  Bd 
xni,  1893,  pp.  790-792. 

A  good  peptone  for  this  purpose  must  be  white,  without 
odor,  entirely  soluble  in  water,  especially  on  warming 
The  water  solution  must  be  clear,  colorless,  neutral  or 
slightly  alkaline,  foamy  on  shaking.  It  must  give  a  vio- 
let reaction  with  Fehling's  solution,  which  does  not 
change  on  boiling.  It  must  give  no  nitrite  reaction  with 
Gnesz  reagent,  and  finally  must  give,  after  about  5  min- 
utes with  Diphenylamine,  a  faint  but  distinct  narrow 
clear  blue  ring. 

('94)-  SCHNEIDER,  PAUL.  Die  Bedeutung  der  Bak- 
terien farbstoffe  fiir  die  Unterscheidung  der 
Arten.  (Inaug.  Diss.)  8vo,  46  pp,  2  Taf, 
Basel,  1894.  Rev.  in  Centralb.  f.  Bakt,  Bd 
xvi,  1894,  p.  633.  See  also  Arbeiten  a.  d. 
bakt.  Institut  Karlsruhe  I,  1894. 

('94)-  MARPMANN.  Zur  Unterscheidung  des  Bacillus 
typhi  abdominalis  vom  Bacillus  coli  com- 
mune. Centralb.  1  Bakt,  Bd.  xvi,  1894,  pp. 
817-820. 


the  first  two. 

('94).  LUNKEWICZ,  M.  Eine  Farbenreaktion  auf  die 
salpetrige  Saure  der  Kulturen  der  Cholera- 
bacillen  und  einiger  anderer  Bakterien 
Centralb.  f.  Bakt,  Bd.  xvi,  1894,  pp.  945-949. 

Describes  the  preparations  of  the  Griess-Ilosvav  re- 
agent. 

('94).  ABEL,  RUDOLF.  Ueber  die  Brauchbarkeit  der 
von  Schild  angegebenen  Formalinprobe  zur 
Differential-Diagnose  des  Typhus  bacillus 
Centralb.  f.  Bakt,  Bd.  xvi,  1894,  PP.  1,041- 
1,046. 

Finds  some  bacteria  more  tolerant  of  formalin  than 
was  stated  by  Schild.  It  is  not  a  satisfactory  method  lor 
separating  typhoid  bacilli  from  B.  coli 


230 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('95).  MACKENSIE,  J.  J.  Opening  discussion  on 
"What  new  methods  can  be  suggested  for 
the  separation  of  bacteria  into  groups,  and 
for  the  identification  of  species."  Jour.  Am. 
Pub.  Health  Asso.,  Oct.,  1895,  Ann.  vol.  xx, 
pp.  419-431- 
Notes  on  synthetic  media. 

('97).  STODDART.    See  xvi. 
('97).  Hiss.    'See  xvi. 

('98).  HOUSTON,  A.  C.  Note  on  four  micro-organ- 
isms isolated  from  the  mud  of  the  river 
Thames,  which  resemble  Bacillus  typhosus. 
Centralb.  f.  Bakt.,  xxiv  Bd.,  1898,  pp.  518- 
525,  colored  diagram. 
('98).  FERMI.  See  xxxvi. 

('98).  ROTHBERGER,  C.  JULIUS.  Differential  diagnos- 
tische  Untersuchungen  mit  gefarbten  Nahr- 
boden.  Centralb.  f.  Bakt.,  xxiv  Bd.,  1898, 
pp.  5I3-SI8. 

With  neutral  red  (Toluidin  red)  in  agar  B.  coli  is  said 
to  cause  a  clearing  of  the  color  and  a  very  decided  fluor- 
escence. B.  typhi  leaves  the  medium  unchanged.  The 
best  method  is  said  to  be  to  add  3  to  4  drops  of  a  concen- 
trated water  solution  to  lo  cc.  of  fluid  agar,  and  then  % 
cc.  of  a  24-hour  old  bouillon  culture.  In  agar  stained 
with  safranin,  B.  coli  reduces  the  color  ;  B.  typhi  does 
not.  Dead  cultures  of  B.  coli  (cultures  heated  2  hours  at 
78°  C.)  did  not  produce  these  changes. 

('98).  PACINOTTI,  G.,  AND  MUNIECKI,  J.  L'albume 
d'uovo  colorito  in  verde-cupo  dal  caffe 
crudo,  come  mezzo  diagnostico  di  svlluppi 
ibatterici.  Gazz.  degli  ospedali  e  della 
dim'che,  1808,  No.  31.  Rev.  in  Centralb.  f. 
Bakt.,  xxv  Bd.,  1899,  p.  257. 

('99.)  ROTHBERGER,  C.  JULIUS.  Differential  diagnos- 
tische  Untersuchungen  mit  gefarbten  Nahr- 
boden.  11,  Mittei-lung.  Centralb.  f.  Bakt., 
xxv  Bd.,  1899,  pp.  15-17  and  69-75. 

Author  tested  the  effect  of  various  bacteria  on  35  anilin 
dves  13  of  which  proved  useful  in  agar  media.  The  loss 
of  color  in  methylen  blue,  safranin,  Toluidin  blue,  Or- 
seille  extract  and  indigo  carmin  is  due  to  reduction  pro- 
cesses. In  case  of  the  indigo  carmin,  the  blue  was  first 
changed  to  dark  green. 

('oo).  SCHEFFLER,  W.  Das  Neutrakot  als  Hilfsmittel 
zur  Diagnose  des  Bacterium  coli.  Centralb. 
f.  Bakt.,  xxvin  Bd.,  1900,  pp.  199-205. 

Author  says  B.  coli  in  neutral  red,  grape  sugar  agar 
gives  regularly  in  24  to  48  hours  a  beautiful  green  fluor- 
escence. He  uses:  fluid  agar,  loo  cc.,  grape  sugar,  0.3 
gram ;  concentrated  watery  solution  of  neutral  red,  ice. 

('oi).  HOELSCHER,  WALTER.  Ueber  die  Differenz  der 
histologischen  Wirkung  von  Tuberkel- 
bacillen  und  anderen  diesen  ahnlichen 
saurefesten  Bacillen  (Grasbacillus  1 1  Moel- 
ler,  Butterbacillus  Petri-Rabinowitsch, 
Thimotheebacillus  Moller).  Munchener 
med.  Wochenschr.,  Bd.  XLVIII,  1901,  pp. 
1,483-1,484- 

('02).  Hiss,  PHILLIP  HANSON,  JR.  New  and  simple 
media  for  the  differentiation  of  the  colonies 
of  typhoid,  colon,  and  allied  bacilli.  The 
Journal  of  Medical  Research,  June,  1902, 
vol.  vin,  pp.  148-167.  Also  a  separate.  2 
plates. 

Author  describes  a  simple  solid  medium  in  which 
buried  colonies  of  the  typhoid  bacillus  send  out  thread- 
formed  radiations,  while  those  of  B.  coli,  etc.,  do  not 
bear  any  fringing  threads.  This  medium,  used  in  Petri- 
dish  poured  plates,  consists  of  distilled  water  1,000,  agar 
15,  gelatin  15,  dextrose  10,  Liebig's  extract  meat  5,  sodium 
chloride  5. 


("02).  FITZ  GERALD,  MABEL  PUREFOY,  AND  DREYER, 
GEORGES.  The  unreliability  of  the  neutral 
red  method,  as  generally  employed  for  the 
differentiation  of  B.  typhosus  and  B  coli. 
Festsksrift  ved  Indvielsen  af  Statens  Serum- 
institut.  Kjobenhavn,  1902,  pp.  1-39. 

('02).  BUXTON,  B.  H.     A  comparative  study  of  the 
bacilli    intermediate   between    B.    coli    coni- 
inunis  and   B.  typhosus.     Jour.    Med.   Res. 
in  (n.  s.),  pp.  201-230,  3  plates. 
Bibliography  of  16  titles. 

("03).  GAGE,  STEPHEN  DEM.,  AND  PHELPS,  EARLE  B. 
Notes  on  B.  coli  and  allied  forms,  with 
special  reference  to  the  neutral-red  'reaction. 
Proceedings  of  the  Thirtieth  Annual  Meet- 
ing, American  Public  Health  Assn.,  New 
Orleans,  La.,  Dec.,  1002.  Pub.  Columbus, 
Ohio,  1903,  vol  xxvin,  pp.  402-412.  Also  a 
separate,  pp.  11. 

('03).  SCHUEDER.  Zum  Nachweis  der  Typhusbak- 
terien  im  Wasser.  Zeitschr.  f.  Hyg.,  Bd. 

XLII,  1003,  PP-  317-326. 
Describes  differential  methods. 

('04).  STOKES,  WILLIAM  ROYAL.     A  simple  test  for 
the  routine  detection  of  the  colon  bacillus  in 
drinking   water.     Jour,   of    Infectious    Dis- 
eases, vol.  i,  1904,  pp.  341-347.     i  plate. 
Neutral  red  reaction. 


XIX.    Aerobism,  Anaerobism. 

(See  also  various  citations  under  XX.) 

('61).  PASTEUR,  Louis.  Animalcules  infuspires 
vivant  sans  gaz  oxygene  libre  et  determinant 
des  fermentation's. — C.  R.  des  se.  de  1'Acad. 
des  sci.,  Paris,  1861,  T.  LII,  pp.  344-347- 

('63).  PASTEUR,  Louis.  Nouvel  exemple  de  fermen- 
tation determinee  par  des  animalcules  in- 
fusoires  pouvant  vivre  sans  oxygene  libre, 
et  en  dehors  de  tout  contact  avec  1'air  de 
I'atmosphere.— C.  R.,  des  se.  -de  1'Acad.  des 
sci.,  Paris,  1863,  T.  LVI,  pp.  416-421. 

('63).  PASTEUR,  Louis.  Recherches  sur  la  putrefac- 
tion. C.  R.  des  se.  de  1'Acad.  des  sci.,  Paris, 
1863,  T.  LVI,  pp.  1,189-1,194. 

"  Je  propose  avec  toute  sorte  de  scrupules  ces  mots 
nouveaux  aerobies  et  anaerobes  pour  indiquer  1'existence 
de  deux  classes  d'6tres  inferieures,  les  uns  incapable  de 
vivre  en  dehors  de  la  presence  du  gaz  oxygene  libre,  les 
autres  pouvant  se  multiplier  a  1'infini  en  dehors  du  con- 
tact de  ce  gaz.'' 

('80).  BUCHNER.      Ueber    die    Lebensfahigkeit    der 

Spailtpilze  bei  fehlenden  Sauerstoff,  1880. 
Not  seen. 

('85).  HESSE,  W.  UND  R.  Ueber  Zuchtung  der 
Bacillen  des  malignen  Oedems.  Deutsche 
med.  Wochenschrift,  1885,  11  Jahrg.,  pp. 
214-215. 

Describes  a  method  of  cultivating  anaerobic  organisms 
by  sowing  them  in  deep  masses  of  solid  media. 

('86).  LIBORIUS,    PAUL.      Beitrage    zur    Kentniss    d. 
Sauerstoffbediirfnisses         der         Bakterien. 
Zeitschr.  f.  Hyg.,  Bd.  i,  1886,  pp.  H5-I77- 
According  to  review  by  Zimmerman  in  Hot.  Centralb. 
Bd.  xxvin,  1886,  p.  198,  author  found  but  little  more  oxy- 
gen in  layers  of  agar  buried  3  cm.  and  over  under  addi- 
tional agar  than  in  vessels  in  which  air  was  expelled  by 
hydrogen,  etc.     He  used  "  Indigotinloesung"  as  a  test. 


AEROBISM,    ANAEROBISM. 


231 


('87).  Roux,  E.  Sur  la  culture  des  microbes 
anaerobies.  Ann.  de  I'lnst.  Pasteur,  Bd.  I, 
1887,  pp.  49-62. 

('88).  BUCHNER,  HANS.  Eine  neue  Methode  zur 
Kultur  anaerober  Mikroorganismen.  Cen- 
tralb.  f.  Bakrt.,  1888,  iv  Bd.,  pp.  149-151. 


('88).  FRAENKEL,  CARL.    Ucber  die  Kultur  anaerober 
Mikroorganismen.     Centralb.  f.  Bakt.,  1888, 
in  Bd.,  pp.  73S-740  and  763-768.  I  fig. 
('89).  FRAENKEL.     See  xxxvm. 

('89).  FRANKLAND,  PERCY.    On  the  influence  of  car- 
bonic anhydride  or  other  gases  on  the  de- 
velopment of  micro-organisms.     Proc.  Roy. 
Soc.,  vol.  XLV,  1889,  pp.  292-301.     See  also 
Zeitsch.  f.  Hygiene,  Bd.  vi,  1889,  pp.  13-22. 
The  other  gases  tested  were  hydrogen,  carbonic  oxide, 
and  nitrons  oxide.    Hydrogen  had  the  least  deleterious 
effect  on  the  organisms   tested,  which  were  B.  pyocy- 
aueus,  Koch's  comma  bacillus,  and  Finkler's  spirillum. 

Coo).  SMITH.    See  xx. 

fyo).  POPOFP.    See  XLvnr. 

('92).  OGATA,  M.  Einfache  Bakterienkultur  mit 
versohiedenen  Gasen.  Centralb.  f.  Bakt., 
xi  Bd.,  1892,  pp.  621-623,  mit  i  fig. 

Ogata's  method,  which  is  essentially  that  previously 
described  by  Heim ,  consists  in  softening  the  upper  part  of 
a  test  tube  in  the  flame  and  drawing  it  out  so  that  the 
tube  consists  of  two  normal  portions  connected  by  a  nar- 
row isthmus.  This  part  should  be  just  below  the  colton 
plug,  that  is,  in  the  upper  one-third  of  the  tube.  A  piece 
of  glass  tubing  is  now  plugged  at  one  end  with  sterile 
cotton,  and  is  drawn  out  into  a  capillary  tube,  which 
must  be  long  enough  to  reach  down  into  the  bottom  of 
the  culture  medium.  The  broken  end  of  this  tube  is  now 
touched  to  the  desired  culture  and  passed  into  the  test 
tube.  The  cotton-plugged  end  of  the  glass  tube  is  now 
attached  to  a  rubber  tube  connected  with  the  gas  appa- 
ratus. After  the  gas  has  bubbled  through  the  medium 
for  a  sufficient  time,  the  capillary  tube  is  removed,  ami 
the  test  tube  is  immediately  sealed  in  the  open  flame  by  a 
further  constriction  of  the  Isthmus  and  a  complete 
removal  of  the  upper  part  of  the  test  tube. 

Fluid  cultures  may  also  be  made  in  such  tubes  by  pre- 
paring the  isthmus  before  the  tubes  are  filled  with  the 
medium. 

('92).  HEIM,     L.       Zur     Originalmittheilung     von 

Ogata:       "Einfache      Bakterienkultur     .mat 

verschiedenen  Gasen."    Bd.  xi,  p.  621.    Cen- 

'tralib.  f.  Bakt.,  xi,  1892,  p.  800. 
('92).  VAN    SENUS,   A.    H.    C.     Zur   Kenntniss    der 

Kultur   anaerober    Bakterien.      Centralb.    f. 

Bakt.,  xii  Bd.,  1892,  pp.  144-145. 
CP3)-  Now,  F.  G.    Die  Kultur  anaerober  Bakterien. 

Centralb.   f.   Bakt,   Bd.   xiv,   1893,  pp.   581- 

600,  2  figs. 

The  "Novyjars"  arc  here  figured  and  described,  and 
49  references  to  literature  are  given  at  the  end  of  the 
article. 

('93).  BKYKRINCK,  M.  W.  Ueber  Atmungsfiguren 
beweglicher  Bakterien.  Centralb.  f.  Bakt., 
Bd.  xiv,  1893,  pp.  827-845,  mit  I  Tafel. 

('93)-  SMITH.    See  xx. 

('94).  NICOLAIER.  Bemerkung  zu  der  Arbeit  von 
Prof.  F.  G.  Novy:  "Die  Kultur  anaerober 
Bakterien"  (Centralb.  f.  Bakt.,  Bd.  xiv, 
1893).  Centralb.  f.  Bakt.,  Bd.  xv,  1894,  p. 
227. 


(94).  SMITH,  THEOBALD.  Further  observations  on 
the  fermentation  tube,  with  special  reference 
to  anaerobiosis,  reduction,  and  gas  produc- 
tion (abstract).  Proc.  Am.  Ass.  A.  Sci., 
42,  Madison  Meeting,  1893,  Salem,  1894, 
p.  261. 

('94).  ENGELMANN,  TH.  W.  Die  Erscheinungsweise 
der  Sauerstoffausscheidung  chromophyllhal- 
tiger  Zellen  im  Licht  bei  Anwendung  der 
Baoterienmethode.  Verhand.  d.  Kon.  Akad. 
van  Wetenschappen  te  Amsterdam  (2  te 
sectie,  deel  in;  No.  11,  1894),  pp.  10,  iv, 
with  a  folded  plate,  also  a  reprint,  17  pp. 
Gives  a  bibliography  of  61  titles.  See  also 
Onderzoekingen  Physiol.  Laborat.  Utrecht, 
iv  Reeks,  in  deel.,  1895. 

('94).  LUBINSKI,  WSEWOLOD.  Zur  Methodik  der 
Kultur  anaerober  Bakterien.  Centralb.  f. 
Bakt.,  Bd.  xvi,  1894,  pp.  20-25,  mit  4  figs. 

('94).  NOVY,  F.  G.  Die  Plattenkultur  anaerober 
Bakterien.  Centralb.  f.  Bakt.,  Bd.  xvi,  1894, 
pp.  566-571,  with  3  figs. 

('94).  ARENS.  Eine  Methode  zur  Plattenkultur  der 
Anaeroben.  Centralb.  f.  Bakt.,  Bd.  xv,  1894, 
pp.  15-17. 

The  author  makes  his  cultures  in  ordinary  exsiccators 
having  ground  glass  covers. 

('95).  SCHMIDT,  AD.  Eine  einfache  Methode  zur 
Zitchtung  anaerober  Kulturen  in  fliissigen 
Nahrboden.  Centralb.  f.  Bakt.,  xvii  Bd., 
!895,  pp.  460-461,  i  fig. 

('95).  BRAATZ,  EGBERT.  Einiges  iiber  die  Anaero- 
biose.  Centralb.  f.  Bakt.,  xvn  Bd.,  1895,  pp 
737-742,  with  i  fig. 

('95).  KEDROWSKI,    W.      Ueber     die     Bedingungen, 
unter  iwelchen  anaerobe  Bakterien  auoh  bei 
Gegcnwart  von  Sauerstoff  existiren  konnen 
Zeitsohr.  f.  Hyg.,  Bd.  xx,  1895,  pp.  358-375- 
They  do   this  best  when  mixed  with  aerobes,  but  the 
absorption  of  the  oxygen  is  not  so  important  as  Pas- 
teur supposed.    The  author  believes  that  the  aerobes  ex- 
crete some  special  substance  favorable  to  the  growth  of 
the  anaerobes.    This  substance  he  did  not  determine. 

('95).  GERSTNER,  R.  Beitrage  zur  Kentniss  obligat- 
anae'rober  Bakterienarten.  Arbeiten  aus 
dem  Bakteriologisohen  Institut  der  Tech- 
nisohen  Hochschule  zu  Carlsruhe,  Bd  i 
1895.  H.  2,  pp.  148-183,  with  2  Taf. 

('96).  KASPAREK,  THEODOR.  Bin  einfacher  Luftab- 
schluss  flussiger  Nahrboden  beim  Kulti- 
vieren  anaerober  Bakterien.  Centralb.  I. 
Bakt.,  xx  Bd.,  1896,  pp.  536-537,  2  figs. 

('96).  DURHAM,  HERBERT  E.  On  a  self-acting  means 
of  cultivating  anaerobic  microbes.  Jour. 
Path,  and  Baot.,  vol.  in,  1896,  pp.  231-236. 

('97).  BECK,  M.  Zur  Ziichtung  anaerober  Kulturen. 
Centralb.  f.  Bakt.,  xxn  Bd.,  1897,  pp.  343- 
345,  with  2  figs. 

('98).  SCHOLTZ,  W.  Ueber  das  Wachstum  anaerober 
Bakterien  bei  ungehindertem  Luftzutritt. 
Zeitschr.  f.  Hyg.,  Bd.  xxvii,  1898,  pp.  132- 
142.  Review  in  Centralb.  f.  Bakt.,  xxrv  Bd., 
1898,  p.  932. 

('98).  FERRAN,  J.  Ueber  das  aerobische  Verhalten 
des  Tetanusbacillus.  Centralb.  f.  Bakt., 
xxrv  Bd.,  1898,  pp.  28-29. 

Thinks  his  experiments  show  tetanus  bacillus  is  not  a 
strict  anaerobe,  but  only  a  facultative  anaerobe. 


232 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


ie  reviewer  has  not  had  that  trouble  with  Novy's  ap- 
itus    complained    of    by    Mr.     Klein.     The    authors 

,    r—+.    ... : ,. ; ,,  ,r    thf>    ru*irn*n  water  WltU 


('98).  KLEIN,  ALEX.  Ein  Apparat  zur  bequemcn 
Herstellung  von  anaeroben  Plattenkulturen. 
Centradb.  f.  Bakt.,  xxiv  Bd.,  1898,  pp.  967- 
971,  2  figs. 

The  rev 

paratus   compiameu   ui    uy    x«i.    «.^....     *- —   -  — 
siphon   arrangement  for  mixing  the  potash  water  with 
the  pyrogallol  after  exhaustion  of  the  air  appears  to  be 
very  good . 

('98)    FESRAN,  J.     Ueber  die  Verwendung  des  Ace- 
tylens  foei  der  Kultur  anaerober  Baktenen. 
Centralb  .f.  Bakt.,  xxiv  Bd.,  1898,  p.  29. 
('98)    ZUPNIK,    LEO.      Ueber    eine    neue    Methode 
anaerober    Zudhtung.      Centralb.    f.    Bakt., 
xxiv  Bd.,  1898,  pp.  267-270,  with  i  fig. 
fq8)    OPRESCU     Zur  Technik  der  Anaerobenkultur. 
Hyg.  Rundschau,  1898,  No.  2.    Rev.  in  Cen- 
tralb. f.  Bakt.,  xxm  Bd.,  1898,  p.  668. 
('98)    UCKE,  ALEXANDER.     Ein  Beitrag  zur  Kenntnis 
der    Anaeroben.      Centralb.    f.    Bakt.,    xxm 
Bd.,  1898,  pp.  996-1,001. 

Co8)    TRENKMANN.     Das  Wachstum  der  anaeroben 
Bakterien.      Centralb.   'f.    Bakt.,    xxm    Bd., 
1898,  pp.  1,038-1,043  and  1,087-1,090. 
('08).  MARPMANN.     Eine   neue   Methode    zur    Her- 
stellung von  anaeroben  Rollglasku.lturen  out 
Gelatine    oder    Agar.      Centralb.    f.    Bakt., 
xxm  Bd.,  1808,  pp.  1,090-1,091. 
('98).  EPSTEIN,    STANISLAUS.     Apparat   zur   Ktiltur 
anaerober    Bakterien.      Centralb.    f.    Bakt., 
xxiv  Bd.,  1898,  pp.  266-267,  i  fig.    , 
('99).  KABRHEL,  GUSTAV.     Zur  Frage  der  Zuchtung 
anaerober    Bakterien.      Centralb.    f.    Bakt., 
xxv  Bd.,  1899,  pp.  55S-S6i,  with  I  fig. 
As  an  oxygen  indicator  author  uses  methylene  blue  in 
sugar  gelatin. 

('99).  SMITH,  THEOBALD.  Some  devices  for  the  cul- 
tivation of  anaerobic  bacteria  in  fluid  media 
•without  the  use  of  inert  gases.  Jour.  Bost. 
Soc.  Med.  Sci.,  1899,  PP-  340-343-  Also  a 
separate,  4  pp. 

Coo)    EPSTEIN,  STANISLAUS.    Em  veremfachtes  Ver- 
fahren    zur   Ziichtung   anaerober    Bakterien 
in  Doppelschalen.    Centralb.  f.  Bakt.,  xxvm 
Bd.,  1900,  p.  443,  with  i  fig. 
Coo).  PETRI.    See  xvii. 
Coo).  KRAUSE.    See  xv. 
Coo).  BULLOCK.    See  xvn. 
Coo).  WRIGHT.    See  xvn. 

('02)  OMELIANSKI,  W.  Ein  einfacher  Apparat  zur 
Kultur  von  Anaeroben  im  Reagenzglase. 
Centralb.  f.  Bakt.,  Abt.  2,  Bd.  vra,  1902,  pp. 


XX.    Fermentations,  Gas-Formation,  Enzymes,  Etc. 

(See  also  XIX  and  XLVIII.) 

('57).  PASTEUR,  L.  Memoire  sur  le  fermentation 
appelee  lactique.  C.  R.  des  se.  de  1'Acad. 
des  sci.,  T.  XLV,  1857,  pp.  913-916. 

('77).  VINES,  S.  H.  On  the  digestive  ferment  of 
Nepenthes.  Journ.  Linn.  Soc.  (Bot),  vol. 
xv,  1877,  pp.  427-43L 

('79).  v.  NAEGELI,  CARL  WILHELM.  Theone  der 
Garung.  Ein  Beitrag  zur  Molecularphysiol- 
ogie.  Miinchen,  1879,  pp.  iv,  156. 

('79).  PRAZMOWSKI,  A.  Zur  Entwicklungsgeschichte 
und  Fermentwirkung  einiger.  Bacterien- 
Arten.  Vorlaufige  Mittheilung.  Bot.  Zeit- 
ung,  Bd.  xxxvn,  No.  26,  col.  409-424,  1879 


('79).  VAN  TiEGHEM,  P.  E.  L.  Sur  la  fermentation  de 
la  cellulose. — Bull,  de  la  Societe  Bot.  de 
France,  1879,  T.  xxvi,  pp.  25-30. 
('82).  MAYER,  ADOLF.  Die  Lehre  von  den  chetnis- 
chen  Fermenten  oder  Enzymologie.  pp.  vi, 
124.  Heidelberg,  1882. 

('82).  BECHAMP,  A.  Sur  les  microzymas  cornine 
cause  de  la  decomposition  de  1'eau  oxygenee 
par  les  tissus  des  animaux  et  des  vegetaux. 
C.  R.  des  se.  de  1'Acad.  des  sci.,  Paris,  T. 
xciv,  pp.  1,653-1,656. 

('82).  BERT,  B.,  ET  REGNARD,  P.  Action  de  1'eau 
oxygenee  sur  les  matieres  organiques  et  les 
fermentations.  C.  R.  des  se.  de  1'Acad.  des 
sci.,  Paris,  T.  xciv,  pp.  1,383-1,386.  Jour, 
de  Pharm.  et  de  Chimie,  T.  vi,  5  serie,  1882, 
pp.  14-17. 

('82).  WORTMANN,  J.  Untersuchuii'gen  uber  das 
diastatische  Ferment  der  Bacterien.  Zeitschr. 
f.  physiol.  chem.,  Bd.  vi,  pp.  287-329,  1882. 
Also  Jour.  Chem.  Soc.,  XLIV,  pp.  930-938, 
1883. 

('82).  MARCANO,  V.  Fermentation  directe  de  la 
fecule.  Mccanisme  de  cette  metamorphose. 
C.  R.  des  se.  de  1'Acad.  des  Sci.,  Paris.  T. 
xcv,  1882,  pp.  856-859. 

('83).  BECHAMP,  A.  Les  microzymas  dans  leurs  rap- 
ports avec  rheterogenie,  I'histogenie,  la 
pliysiologie  et  la  pathologic,  pp.  xxxvni,  992, 
with  3  pi.  Paris,  1883. 

Prior  to  1883  Bechnmp  isolated  from   yeast  an  enzyme 
capable  of  inverting  cane  sugar.     See  pp.  69  et  seq. 

('83).  VIGNA,  A.  Ueber  Bakteriengahrung  des 
Glycerins.  Ber.  d.  deutsch.  chem.  Gesells., 
xvi,  pp.  1,438-1,439;  Gazz.  Ghim.  Ital.,  vol. 
xm,  pp.  293-296,  1883. 

('83).  TAPPEINER,  H.  Ueber  Cellulosegahrungen. 
Ber.  d.  deutsch.  chem.  Gesells.,  Bd.  xvi, 
1883,  pp.  1,734-1,740. 

('83).  TAPPEINER,  H.  Ueber  die  Sumpfgasgahrun» 
im  Schlamme  der  Teiche,  Siimpfe  und 
Kloaken.  Ber.  d.  deutsoh.  chem.  Gesells., 
Bd.  xvi,  1883,  pp.  1,740-1,744. 

('86).  HOPPE-SEYLER,  FELIX.  Ueber  Gahrung  der 
Cellulose  mit  Bildung  von  Mebhan  und 
Kohlensaure.  Zeitschr.  f.  physiolog.  Chemie., 
1886,  Bd.  x,  pp.  201-217  and  401-440. 

('89).  KRAMER,  ERNST.  Studien  fiber  schleimige 
Gahrung.  Sitzungsb.  d.  k.  Ak.  d.  Wissen- 
schaften.  Math.-naturw.  Classe,  Bd.  xcvni, 
1889,  iv  Hft,  Abt.  ii  b.  Chemie,  pp.  358-396. 
Wien. 
An  interesting  paper. 

('89).  FRANKLAND,  PERCY  P.,  AND  Fox,  J.  J.  On  a 
pure  fermentation  of  mannite  and  glycerine. 
Proc.  Royal  Soc.,  Lond.,  1889,  vol.  XLVI,  pp. 
345-357- 

('89).  CURTMAN,  CHAS.  O.  Naoh-weis  der  Glycose 
durch  Safranin.  Pharmaceutische  Runds- 
chau, Bd.  vn,  1889,  p.  132. 

('89).  TISCHUTKIN,  N.  Die  Rolle  der  Bakterien  bei 
der  Veranderung  der  Eivveissstoffe  auf  den 
Blattern  von  Pinguicula.  Ber.  d.  deutsch. 
'bot.  Gessellsch.,  Bd.  vn,  1889,  pp.  346-355- 

('90).  SCLAVO,  ACHILLE,  UND  Gosio,  B.  Ueber  eine 
neue  Garung  der  Starke.  Staz.  sperim. 
agrar.  ital.,  1890,  vol.  xix,  p.  540. 


FERMENTATIONS,    GAS-FORMATION,    ENZYMES,    ETC. 


233 


('90).  REINITZKR,  FR.  Uober  die  wahre  Natur  des 
Gummifermentes.  Zeitschr.  S.  physiol. 
Ohcmie,  Bd.  xiv,  1890,  pp.  453-470. 

This  author  states  that  Wiesner's  ferment,  which 
changes  cellulose  into  gum  and  slime,  does  not  exist. 
Nickel,  of  Berlin,  has  come  to  the  same  conclusion  inde- 
pendently. (See  Bot.  Centralb.,  1890,  Bd.  xi.in,  p.  118). 
The  "orciu  salzsiiure  "  reaction  of  Reichl  is  a  furfurol 
reaction. 

Coo).  SMITH,  THEOBALD.     Das  Gahrungskolbchen  in 

der    Bakteriologie.     Centralb.    f.    Bakt.,   vn 

Bd.,  1890,  pp.  502-506,  mit  i  Abbildung. 

('90).  SMITH,  THEOBALD.    Einige  Bemerkungen  fiber 

Satire-    und    Alkalibildung    bei    Bakterien. 

Centralb.  f.  Bakt.,  Bd.  vm,  1890,  pp.  389-391. 

('90).  FERMI,  CLAUDIO.    Die  Leim  und  Fibrin  losen- 

den    und    die    diastatischen    Fermente    der 

Mikroorganismen.     Centralb.    f.    Bakt.,   vii 

Bd.,  1890,  pp.  469-474. 

('90).  DUBOIS,  R.  Sur  le  pretendu  pouvoir  digestif 
du  liquide  de  1'urne  des  Nepenthes.  C.  R. 
des  se.  de  I'Acad.  des  Sci.  T.  cxi,  1890,  pp. 
315-317. 

Maintains  that  the  observed  results  are  not  due  to  any 
pepsin-like  body  secreted  by  the  plant,  but  to  the  action 
of  intruding  bacteria,  and  concludes  that  the  Nepenthes 
are  not  carnivorous. 

('91).  HERY.  ,Sur  une  fermentation  visqueuse  de 
1'encre.— Annales  de  Micrographie,  1891,  T. 
rv,  pp.  13-21. 

(91).  FERMI,  CLAUDIO.  Wekere  Untersuchungen 
iiber  die  itryptischen  Enzyme  der  Mikroor- 
ganismen. Centralb.  f.  Bakt.,  x  Bd.,  1891, 
pp.  401-408. 

('91).  RITSERT.  Bakteriologische  Untersuehungen 
iiber  das  Schleimigwerden  der  Infusa.  Ber. 
d.  pharm.  Gesellsch.,  I  Bd.,  1891,  pp.  389- 
309.  Rev.  in  Centralb.  f.  Bakt.,  1892,  xi, 
PP-  730-733. 

('91).  SUCHSLAND,  E.  Ueber  Tabaksfermentationen. 
Ber.  d.  d.  bot.  Gesellsch.,  Bd.  ix,  1891,  pp. 
79-8 1. 

('91).  BEYERINCK,  M.  W.  Verfahren  zum  Nachwcise 
der  Saureahsonderung  bei  Mikrobien.  Cen- 
tralb. f.  Bakt.,  ix  Bd.,  1891,  pp.  781-786. 

This  method  consists  in  using  with  gelatin  or  agar,  for 
plate-cultures,  etc.,  a  solid  opaque  substance  which  is 
converted  into  a  soluble,  transparent  substance  in  the 
presence  of  acids.  For  this  purpose  Beyerinck  adds  to 
the  culture- medium  a  small  amount  of  water  in  which 
a  very  finely  divided  carbonate  of  lime  has  been  shaken 
up.  This  makes  a  white  opaqne  plate.  Colonies  which 
secrete  acids  are  soon  surrounded  by  a  transparent  dif- 
fusion field.  Other  carbonates  may  be  used,  e.  g  ,  zinc 
carbonate.  This  method  may  also  be  used  for  the  demon- 
stration of  alkali  production. 

('91).  TISCHUTKJN,  N.  P.  Ueber  die  Rolle  der 
Mikroorganismen  bei  der  Ernahrttng  der 
insektenfressenden  Pflanzen.  Arbeit,  d.  St. 
Petersburg  naturf.  Gesell.,  vol.  for  1890, 
Sect.  d.  Bot.  pp.  33-37,  St.  Petersburg,  1891. 
Abs.  in  Bot.  Centralb.,  Bd.  L,  1892,  p.  304. 

C'jjt).  CONN,  H.  W.  Isolirung  eines  "Lab"  fer- 
mentes  aus  Bakterienkulturen.  Centralb.  f. 
Bakt.,  Bd.  xn,  1892,  pp.  223-227. 

('92?).  FRANKLAND  AND  FREW.  The  fermentation  of 
calcium  glycerate  by  the  Bacillus  ethaceticus. 
Trans.  Roy.  Ohem.  Soc.,  1891.  Rev.  in 
Centralb.  f.  Bakt..  xn  Bd.,  1892,  p.  724. 

('92).  FRANKLAND,  P.,  AND  MACGREGOR,  J.  Fermen- 
tation of  arabinoM  with  the  Bacillus  etha- 
ceticus. Trans.  Chem.  Soc.,  1892.  Rev.  in 
Centralb.  f.  Bakt.,  xn  Bd.,  1892,  p.  725. 

("92).  FRANKLAND,  P.  P.,  AND  FREW,  W.  A  pure 
fermentation  of  mannitol  and  dulcitol.  Trans. 
Chem.  Soc.,  1892,  pp.  254-277. 


('92).  FRANKLAND,  P.  P.,  AND  LUMSDEN,  J.  S.  De- 
composition of  mannitol  and  dextrose  by  the 
Bacillus  ethaceticus.  Trans.  Chem.  Soc., 

1892,  pp.  432-444. 

('92).  FERMI,   CLAUDIO.     Beitrag  zum   Studium  der 
von    den    Mikroorganismen    abgesonderten 
diastatischen  und  Inversionsfermente.    Cen- 
tralb. i.  Bakt.,  xn  Bd.,  1892,  pp.  713-715. 
Of  62  bacteria  investigated,  20  were  acid  producers,  2  in- 
verted cane  sugar,  24  developed  a  proteolytic  enzym,  20  a 
diastasic  ferment. 

('92).  TISCHUTKIN,  N.  Ueber  die  Rolle  der  Micro- 
organismen  bei  der  Erna/hrung  insekten- 
fressender  Pflanzen.  Acta  Horti  Petro- 
politanie,  Bd.  xn,  St.  Petersburg,  1892,  pp. 
1-19.  Abstract  in  Bot.  Centralb.,  Bd.  LIU, 

1893,  p.  322. 

('92).  FRANKLAND,  P.  P.     Cantor  lectures  on  recent 
contributions  to  the  chemistry  of  bacteriol- 
ogy of  the  fermentation  industries,  gr.  8vo., 
31  pp.    London,  W.  S.  Trounce,  1892. 
Not  seen. 

('93).  HILDEBRANDT,  H.  Weiteres  fiber  hydrolytische 
Fermente,  etc.  Arch.  f.  path.  Anat.  u. 
Physiol,  Bd.  cxxxi,  1893,  pp.  5-39. 

('93)-  GOEBEL.  Pflanzenbiologische  Sehilderungen  ii, 
1893,  p.  186. 

Criticises  views  of  Duhois  and  Tischutkin. 

('93).  HAPP.    Bakteriologische  und  chemisohe  Unter- 
suchungen  fiber   die   sohleimige   Garung,  p. 
31,  these  inaugurate.    Berlin,  1893. 
Not  seen.    Describes  Micrococcus  gummosiis. 

('93).  CAVAZZANI,  EMIL.  Zur  Kenntnis  der  diasta- 
tischen Wirkung  der  Bakterien.  Centralb. 
f.  Bakt.,  Bd.  xm,  1893,  pp.  587-589. 
('93).  PER£,  A.  Sur  la  formation  des  acides  lactiques 
isomeriques  par  1'action  des  microbes  sur 
les  substances  hydrocarbonees.  Ann.  de 
1'Inst.  Pasteur,  T.  vii,  1893,  pp.  737-750. 

('93).  HESSE,  W.  Ueber  die  gasformigen  Stoff- 
wechselproducte  beim  Waohsthum  der  Bak- 
terien. Zeitschr.  f.  Hyg.,  Bd.  xv,  1893,  pp. 
17-37- 

('93).  SMITH,  THEOBALD.  The  fermentation  tube 
with  special  reference  to  anaerobiosis  and 
gas  production  among  bacteria.  Wilder 
Quarter  Century  Book,  Ithaca,  N.  Y.,  1893, 
pp.  187-232. 

('93).  FRANKLAND,  P.,  AND  MACGREGOR,  J.  Sarco- 
lactic  acid  obtained  by  the  fermentation  of 
inactive  Lactic  acid.  Trans.  Chem.  Soc., 
London,  1893,  pp.  1,028-1,035. 

('93).  GRIMBERT.  L.  Fermentation  anaerobie  produite 
par  le  Bacillus  prthobutylicus,  ses  variations 
sous  certaines  influences  biologiques.  Ann. 
de  1'Inst.  Pasteur,  T.  vii,  1893,  pp.  353-402. 

('93).  GREEN,  J.  R.  On  Vegetable  Ferments.  An- 
nals of  Botany,  1893.  Vol.  vii,  pp.  83-137. 

('94).  FERMI,  CLAUDIO,  AND  PERNOSSI,  LEONE.  Ueber 
die  Enzyme.  Centralb.  f.  Bakt.,  Bd.  XV, 
1894,  pp.  229-234. 

('94).  FERMI  UND  PERNOSSI.  Ueber  die  Enzyme. 
Zeitschr.  f.  Hyg.,  Bd.  xvin,  1894,  pp.  83-127. 

Studies  of  the  action  of  heat,  light,  and  various  chemi- 
cal substances  on  various  enzymes. 

('94).  WOOD,  JOSEPH  T.  Fermentation  in  the  leather 
industry.  Journal  of  the  Society  of  Chemi- 
cal Industry,  1894,  vol.  xni,  pp.  218-222. 


234 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('94).  GOTTSTEIN,  ADOLF.  Ueber  die  Zerlegung  des 
Wasserstoffsuperoxyds  durch  die  Zellen, 
imit  Bemerkungen  iiber  eine  makroskopische 
Reaktion  fiir  Bakterien.  Virchow's  Archiv., 
Bd.  cxxxui,  p.  296.  Rev.  in  Centralb.  f. 
Bakt.,  Bd.  xvi,  1894,  pp.  518-519- 

('94).  TIMPE,  HERMANN.  Ueber  die  Beziehungen  der 
Phosphate  und  des  Kaseins  zur  Milchsaure- 
garung.  Die  landwirtschaft.  Versuchsstat., 
Bd.  XLII,  1894,  pp.  223-238. 

('94).  LASER,  HUGO.  Die  makroskopische  Wasser- 
untersuchung  durch  Anwendung  von  Was- 
serstoffsuperoxyd.  Centralb.  f.  Bakt.,  Bd. 
xvi,  1894,  pp.  180-182. 

This  substance,  which  causes  liberation  of  oxygen  from 
H2O2,  is  said  to  be  nuclein.  Author  failed  to  confirm 
Gottstein's  results.  (Virchow's  Archiv.,  Bd.  cxxxui, 
Heft  2.) 

('94).  GREEN,  J.  R.  The  influence  of  light  on  dias- 
tase. Abstr.  of  a  paper  read  before  the 
British  Assoc.  at  Oxford,  Aug.,  1894.  Ann. 
of  Bot.,  vol.  vin,  1894,  pp.  370-373- 
('94).  Gosio,  B.  Ueber  Links-Milchsaure  bildende 
Vibrionen.  Arch.  f.  Hyg.,  Bd.  xxi,  1894, 
pp.  114-122. 

('94).    MACFAYDEN   AND   BLAXALL.      See   XXXIV. 

('95).  BAIER,  EDUARD.  Ueber  Buttersauregalirung. 
Centralb.  f.  Bakt.,  2  Abt.,  Bd.  I,  1895,  pp. 
84-87  and  118-120. 

('95).  OMELIANSKI,  V.  Sur  la  fermentation  de  la 
cellulose.  C.  R.  des  se.  de  1'Acad.  des  sci., 
T.  cxxi,  1895,  pp.  653-655. 

('95)-  JOERGENSEN,  ALFRED.  Les  microorganismes  de 
la  fermentation.  Traduit  par  Paul  Freund. 
8vo.,  318  pp.,  56  illst.,  Paris  (Soc. 
d'editions  scientifiques),  1895.  Micro- 
organisms and  Fermentation.  English  trans- 
lation bv  Alex.  K.  Miller  and  A.  E.  Lenn- 
holm.  Third  edition  completely  revised,  with 
83  illustrations,  pp.  xm,  318.  London, 
Macmillan  &  Co.,  Ltd.  New  York,  The 
Macmillan  Co.,  1900.  There  is  also  a  Ger- 
man -translation  which  has  reached  the  3d 
ed.  Berlin,  1892. 
Orig.,  in  Danish,  not  seen. 

('95).  Gosio,  B.  Zersetzungen  zuckerhaltigen 
Nahrmateriales  durch  den  Vibrio  choleras 
asiaticas  Koch.  Arch.  f.  Hyg.,  Bd.  xxn, 
1895,  pp.  1-27. 

('95).  FERMI  UND  MONTESANO.    Die  von  den  Mikro- 
ben    bedingte    Inversion    des    Rohrzuckers. 
Centralb.  f.  Bakt.,  2  Abt.,  Bd.  I,  1895,  No. 
13-14,  pp.  482-487,  No.  15-16,  pp.  542-556. 
The   author  used  Nylander's   and    Rubner-Penzoldt's 
reactions.     Fehling's  solution  was  not  used,  because  it  is 
not  to  be  trusted  in  the  presence  of  albumen. 

('95).  BEYERINCK,  M.  W.  Ueber  Nachweis  und  Ver- 
breitung  der  Glukase,  das  Enzym  der  Mal- 
tose. Centralb.  f.  Bakt.,  2  Abt.,  Bd.  I,  1895, 
pp.  221-229,  pp.  265-271,  and  pp.  329-342. 

('96).  BOURQUELOT,  EMIL.  Les  ferments  solubles 
(diastases-enzymes),  pp.  yin,  220.  8vo. 
Paris,  Societe  d'editions  scientifiques,  1896. 

('96).  PFEFFER,  W.  Ueber  regulatorisehe  Bildung 
von  Diastase.  Ber.  u.  d.  Verhandl.  d.  K. 
saoh.  Gesell'Sch.  d.  Wissenschaften  zu  Leip- 
zig. Mathematisch-Physik.  Classe,  1896,  pp. 
513-518.  Also  a  separate. 

('97).  BUCHNER,  H.  Die  Bedeutung  der  activen 
loslichen  Zellprodukte  fur  den  Ohemismus 
der  Zelle.  Miinchen.  med.  Wochenschr., 
1897,  No.  12,  pp.  209-302. 


('97).  BUCHNER,  ED.  Alkoholische  Giiruiig  ohnc 
Hefezellen.  Ber.  d.  deutsch.  chem.  Ge- 
sellsoh.,  1897,  Jahrg.  xxx,  pp.  117-124  and 
1,110-1,113.  See  also  Ibid.,  Jahrg.  xxxi,  p. 

568-574- 

"  Eine  Trenmmg  der  Gahrwirkuug  von  den  lebendeu 
Hefezellen  ist  bisher  nicht  gelungen."     (XXX,  p.  117.) 

(*97)-  VINES,  S.  H.  The  proteolytic  enzyme  of 
Nepenthes.  Annals  of  Botany,  vol.  xi,  1897, 

PP.  563-584- 

('98).  PRESCOTT,  S.  C.,  AND  UNDERWOOD,  W.  LYMAN. 
Contributions  to  our  knowledge  of  micro- 
organisms and  sterilizing  processes  in  the 
canning  industries.  II  The  souring  of 
canned  sweet  corn.  Tech.  Quarterly,  vol. 
xi,  No.  i,  1898,  6  plates.  Also  a  separate, 
30  pp. 

('99).  OMELIANSKI,  V.  Sur  la  fermentation  de  la 
cellulose.  Arch,  des  sci.  biol.  publiees  par 
1'inst.  imp.  de  med.  exper.  a  St.  Petersbourg, 
T.  vii,  pp.  411-434.  i  heliotype  plate. 

('99).  FERMI,  CLAUDIO,  AND  BUSCAGLIONI.  Die  pno- 
teolytischen  Enzyme  im  Pflanzenreiche. 
Centralb.  f.  Bakt.,  2te  Abt.,  Bd.  v,  1899,  No. 
i,  pp.  24-27;  No.  2,  pp.  63-66;  No.  3,  pp. 
91-95;  No.  4,  pp.  125-134;  and  No.  5,  pp. 
145-158. 

('99).  DUCLAUX,  E.  Traite  de  microbiologie.  Tome 
ii.  Diastases,  toxines  et  venins.  Paris, 
Masson  et  Cie,  1809,  pp.  ill,  768,  large  8vo. 
The  best  treatise  in  French. 

('99).  SACHAROFF,  N.  Einige  erganzende  Angaben 
zur  Mitteilung:  "Ueber  den  Ohemismus  der 
Wirkung  der  Enzyme  und  der  bakterioiden 
Stoffe."  Centralb.  f.  Bakt.,  xxv  Bd.,  1899, 
PP-  346-350. 

('99).  BULLEK,  A.  H.  R.  Die  Wirkung  von  Bak- 
iterien  auf  tote  Zellen.  Inaugural.  Disserta- 
tion. Univ.  of  Leipsic,  pp.  47.  Leipsic,  1899. 

('99).  BREDIG,  G.,  UND  MUELLER  v.  BERNECK,  R.  Ueber 
Platinkatalyse  und  die  chemiische  Dynamik 
des  Wassers'toffsuperoxyds.  Zeitschr.  fiir 
Physikalische  Chemie.,  Bd.  xxxi,  1899,  pp. 
258-353,  3  'text  figs. 

In  many  ways  platinum  black  behaves  like  an  enzyme. 

('99).  BUCHNER,    E.,   UND   RAPP,    R.     Alcoholisohe 

Gahrung  ohne  Hefezellen.     Ber.  d.  deutsch. 

chem.  Gesellsch.,  Jahrg.  xxx   (1897),  2,668; 

Jahrg.  xxxi  (1898),  209,  1,084,  1,090,  i,S3i; 

Jahrg.  xxxn  (1899),  127-137. 
('99).  GREEN,   J.    REYNOLDS.     The   soluble    ferments 

and  fermentation,    pp.  xm,  480.    Cambridge 

(England),  at  the  University   Press,   1809. 

2d  ed.,  1901,  pp.  XV,  512,  'with  a  bibliography 

of  896  titles. 

An  excellent  book,  and  the  only  complete,  authorita- 
tive one  in  English. 

Coo).  DUCLAUX,  E.  Traite  de  microbiologie.  Tome 
in.  Fermentation  alcoolique.  Paris,  1900, 
pp.  in,  760,  84  text  figures,  8vo.  Masson 
et  Cie. 

Coo).  TURRO,  R.  Zur  Bakterienverdauung.  Cen- 
tralb. f.  Bakt.,  xxvin  Bd.,  1900,  pp.  173-177- 

('oo).  MORGENROTH,  J.  Zur  Kenntnis  der  Labenzyme 
und  ihrer  Antikorper.  Centralb.  f.  Bakt., 
xxvn  Bd.,  1900,  pp.  721-724. 

COT).  SCHOENE,  ALBERT.  Die  Mikroorganismen  in 
den  Saften  der  Zuckerfabriken.  Berlin,  Zs. 
Ver.  D.  Zuckerind.,  Bd.  LI,  1901,  techn.  Tl, 
pp.  453-468. 


FERMENTATIONS,  ENZYMES;  PTOMAINES,  TOXINS,  ETC. 


235 


('oi).  BOUHFARD,  A.  Les  maladies  microbiennes  des 
vins.  Fermentation  alcoolique;  maladies 
microbiennes ;  carre  des  vins ;  hygiene  des 
vins;  traitement  des  vins  malades.  Nancy, 

1901,  I2mo.,  avec  6  planches  et  fig. 

Not  seen. 

('oi).  DUCLAUX,  E.  Traite  de  microbiologie.  Tome 
IV.  Fermentations  variees  des  diverses  sub- 
stances ternaires.  Paris,  Masson  et  Cie., 
loor,  pp.  in,  768,  45  text  figures. 

Valuable. 

('oi).  GKSSAKD,  C.     fitodes  sur  la  tyrosinase.     Ann. 

de  I'lnst.  Pasteur,  T.  xv,  1901,  pp.  593-614. 
('oi).  BEIJERINCK,  M.  W.    Anhaufungsversuche  mil 

Ureumbakterien.        Ureumspaltung      duroh 

Urease  und  durch  Katabolismus.     Centralb. 

f.  Bakt.,  Bd.  vn,  1901,  pp.  33-61,  with  I  plate 

and  4  figures. 
("02).  RoLLY.      Weiterer    Beitrag    zur    Alkali-    und 

Saureproduktion   der    Bakterien.     Arch.    f. 

Hyg.,  Bd.  XLI,  1902,  pp.  406-412. 
('02).  SCHREIBER,      KARL.        Fettzersetzung      durch 

Mikroorgani&men.     Arch.  f.  Hyg.,  Bd.  XLI, 

1902,  pp.  328-347. 

('02).  SMITH,  R.  GREIG.  The  gum  fermentation  of 
sugar  cane  juice.  Proc.  Linnean  Soc.,  N. 
So.  Wales,  for  the  year  1901,  Sydney,  1902. 
Vol.  xxvi,  Pt.  i,  pp.  589-625,  i  plate.  Also 
a  separate. 

('02).  SMITH,  R.  GREIG.  The  deterioration  of  raw 
and  refined  sugar  crystals  in  bulk.  The  acid 
fermentation  of  raw  sugar  crystals.  Proc. 
Linn.  Soc.  of  New  South  Wales,  for  1901, 
Pt.  4,  pp.  674-683,  684,  Sydney,  1902.  Also 
a  separate  (issued  May  20,  1902). 

('02).  OMKUANSKI,  W.  Sur  la  fermentation  fpr- 
menique  de  la  cellulose.  Arch,  des  sci.  biol. 
publiees  par  .1'inst.  imp.  de  med.  exper.  a 
St.  Petersbourg,  T.  ix,  1902,  pp.  251-278,  I 
heliotype  plate. 

('02).  EFFRONT,  JEAN.  Enzymes  and  their  applica- 
tions. English  translation  by  Samuel  C.  Pres- 
cobt,  vol  i.  The  enzymes  of  the  carbohy- 
drates. The  oxidases.  New  York,  John 
Wiley  i&  Sons ;  London,  Chapman  &  Hall, 
Ltd.,  1902,  pp.  xi,  322. 

('02).  SAWAMURA,  S.  On  the  liquefaction  of  mannan 
by  microbes.  Bulletin  of  the  College  of 
Agriculture,  Tokyo  Imperial  University,  vol. 
v,  No.  2,  1902,  pp.  259-262.  Also  a  separate. 

('02).  WEISS,  RICHARD.  Uber  die  Baktcrienflora  der 
Sauern  Gahrung  einiger  Nahrungs-und 
Genussmittl.  Arb.  a.  d.  Bact.  Institut  der 
tech.  Hochschule  zu  Karlsruhe,  11  Bd.,  3 
Heft,  1902,  pp.  163-269. 

Forty-eight  new  species  are  described  :  Streptococcus 
citreus,  S.  maximus,  Micrococcus  pulcher,  M.  regularis, 
M.  irregularis,  M.  umbilicatus,  M.  minimus,  M.  gummo- 
sus,  M.  mucitaginens,  M.  vulgaris,  M.  plliformis,  M. 


II.  eminans,  B.  ventricosus,  B.  citricus 
amygdaloides,  B.  robustus,  B.  tuberosus 
I!,  fungosiis,  B.  flavescens,  B.  grnciles 
variosum,  B.  spinosum,  B.  crenatnm,  B 
brevissimum,  B.  raiuificans,  B.  gibbosnm 
B.  squanintinii,  B.  unifonne,  B.  insnlsum 
B.  gramilosum,  Pseudomonas  lactica,  Ps 


B.  odoratus,  B 
B.   globulosus 
ens,    Bacteriun 
plicativum,  B 
B.  gracillimum 
B.  subcitrinuni 
Listed. 

('02).  GRAN,  H.  H.  Studien  iiber  Meeresbakterien. 
2.  Ueber  die  Hydrolyse  des  Agar-Agars 
durch  ein  neues  Enzym,  die  Gelase.  Bergens 
Mus.  Aarb.,  1902,  No.  2,  p.  16. 


('03).  KELLERMAN,  KARL.  The  effects  of  various 
chemical  agents  upon  the  starch-converting 
power  of  taka  diastase.  Bulletin  Torrey 
Botanical  Club,  vol.  xxx,  1903,  pp.  56-70. 
Also  a  separate. 

('03).  VINES,  S.  H.  Proteolytic  enzymes  in  plants. 
Annals  of  Botany,  vol.  XVH,  Jan.,  1903,  pp. 
237-264. 

('03).  SMITH,  R.  GREIG.  A  gum  (levan)  bacterium 
from  a  saccharine  exudate  of  Eucalyptus 
Stuartiana.  Proc.  Linnean  Soc.,  N.  So. 
Wales,  vol.  xxvn,  for  the  year  1902,  pp.  230- 
236.  i  plate.  Sydney,  ioo2-'o3. 

('03).  LIPPMANN,  E.  O.  VON.  Zur  Nomenklatur  der 
Enzyme.  Berichte  d.  deutsch.  Chem.  Gesell- 
schaft,  1903,  Bd.  xxxvi,  p.  331. 


XXI.     Ptomaines,  Toxins,  Antitoxins,  Serums, 
Phagocytosis,  etc. 

('86).  BKIECER.       Untersuchungen     iiber     Ptomaine. 

Berlin,    1886.      Traduotion    par    Roussy    et 

Winter  as  Microbes,  ptomaines  et  maladies. 

Paris,  1887,  pp.  xii,  235. 
('87).  METCHNIKOFF,  ELIE.    Sur  la  lutte  des  cellules 

de      1'organisme      centre      1'invasion      des 

microbes.     Ann.    de    I'lnst.    Pasteur,   T.    i, 

1887,  pp.  321-336. 
('88).  NUTTALL,    GEO.      Experiments    fiber   die   bac- 

terienfeindlichen     Einfluss    des    thierisohen 

Korpers.    Zeitschr.  f.  Hyg.,  Bd.  iv,  1888,  pp. 

353-394- 
('88).  GAERTNER.      Ueber    die    Fleischvergiftung   in 

Krankenhauser  und  den  Erreger  derselben 

Jena,  1888. 

Not  seen. 

('89).  NISSEN,  F.  Zur  Kenntniss  der  Bacterien- 
vernichtenden  Eigensohaft  des  Blutes. 
Zeitsohr.  f.  Hyg.,  Bd.  vi,  1889,  pp.  487-520. 

('89).  BUCHNER,  H.  Ueber  die  baoterientodtende 
Wirkung  des  zellenfreien  Blutserums.  Cen- 
tralb. f.  Bakt.,  Bd.  v,  1889,  pp.  817-823;  and 
Bd.  vi,  1889,  pp.  i-n. 

('90).  BRIEGER,  L.,  u.  FRAENKEL,  CARL.  Untersuchun- 
gen iiber  Bacteriengifte.  Berl.  klin.  Woch- 
enschr.,  Bd.  xxvn,  1890,  No.  n,  pp.  241-246; 
No.  12,  pp.  268-271. 

('90).  LEHMANN.  Ueber  die  pilztodtende  Wirkung 
des  frisohen  Harns  des  gesunden  Menschen. 
Centralb.  f.  Bakt.,  Bd.  vn,  1890,  pp.  457-460. 

("90).  JACQUEMART,  F.  Les  ptomaines,  histoire  et 
caracteres  chimiques,  memoire  oouronne  par 
la  Societe  royale  des  sciences  medicaks  eit 
naturelles  de  Bruxelles.  Jour,  de  med.  de 
chir.  et  de  pharm.,  Bruxell«s,  1890,  No.  18. 
Rev.  in  Centralb.  f.  Bakt.,  ix  Bd.,  1891,  pp. 
107-110. 

("90).  CARBONE,  TITO.  Ueber  die  von  Proteus  vul- 
garis  erzeugten  Gifte.  Centralb.  f.  Bakt., 
1800,  Bd.  vm,  pp.  768-773. 

('90).  LEWANDOWSKI,  A.  Ueber  Indol-  und  Phenol- 
Wldung  durch  Bakterien.  Deutsche  mediz. 
Wochenschrft,  Bd.  xvi,  1890,  p.  1,186. 

('90).  GABRITCHEVSKY,  G.  .Sur  les  proprietes  chimio- 
tactiques  des  leucocytes.  Ann.  de  I'lnst. 
Pasteur,  T.  iv,  1890,  pp.  346-362. 

('91).  OGATA,  M.  Ueber  die  bacterienfeindliche 
Substanz  des  Blutes.  Centralb.  f.  Bakt., 
Bd.  ix,  1891,  pp.  597-602. 


236 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('gi ).  TRAPEZNIKOFF.  Du  sort  des  spores  de 
microbes  dans  1'organisme  animal.  Ann.  de 
•1'Inst.  Pasteur,  T.  v,  1891,  pp.  362-394,  2 
plates. 

('94).  EHRLICH,  P.,  UND  WASSERMANN,  A.  Ueber 
die  Gewinnung  der  Diphtherie-antitoxine 
aus  Blutserum  und  Milch  iinmunisirter 
Thiere.  Zeitsch.  f.  Hyg.,  Bd.  xvm,  1894, 
pp.  239-250. 

('9=0  BRIEGER,  L.  Wetter*  Erfahrungen  uber  Bak- 
teriengifte.  Zeitsch.  f.  Hyg.,  Bd.  xix,  1895, 

pp.    IOI-II2. 

('96)  BRIEGER  UND  BOER.  Ueber  Antitoxine  und 
Toxine.  Zeitsch.  f.  Hyg.,  1896,  Bd.  xxi,  pp. 
259-268. 

('96).  GAUTIER,  ARM  AND.  Les  toxines  microbiennes 
et  animates,  1896,  pp.  vn,  617.  Soc.  d'edi- 
tions  scientifiques,  Paris. 

('96).  EHRLICH,  P.  Die  staatliche  controle  des  Diph- 
theric serums.  Berl.  klin.  Wochenschr.,  Bd. 
xxxm,  1806,  pp.  441-443- 

('96).  VAUGHAN,  VICTOR  C.,  AND  Now,  FREDR.  G. 
Ptomains,  leucomains,  toxins,  and  anti- 
toxins. 3d  ed.  Lea  Bros.  &  Co.,  Philadel- 
phia and  New  York,  1896,  pp.  x,  604. 

Contains  a  bibliography  of  several  hundred  titles.  The 
4th  ed.  was  issued  in  1902,  with  the  following  title:  Cellu- 
lar toxins,  or  the  chemical  factors  in  the  causation  of 
disease.  Same  publishers.  Pp.  vm.  495- 

('96).  METCHNIKOFF,  EL.,  Roux,  E.,  ET  TAURELLI- 
SALIMBENI.  Toxine  et  antitoxine  cholerique. 
Ann.  de  1'Inst.  Pasteur,  T.  x,  1896,  pp.  257- 
282. 

('97).  SMITH,  THEOBALD.  A  modification  of  the 
method  for  determining  the  production  of 
indol  by  bacteria.  Jour.  Exper.  Med.,  vol. 
n,  1897,  pp.  543-547- 

Coo).  BORDET,  J.  Les  serums  (hemolytiques,  leurs 
antitoxines  et  les  (theories  des  serums  cytoly- 
tiques. Ann.  de  1'Inst.  Pasteur,  1900,  T. 
xiv,  pp.  257-296. 

Coo).  METCHNIKOFF,  E.  Sur  les  cytotoxmes.  Ann. 
de  1'Inst.  Pasteur,  T.  xrv,  pp.  36p-377,  1900. 

Coo).  FISCHER,  ALFRED.  Die  Empfindliohkeit  der 
Bakterienzelle  und  das  bakterieide  Serum. 
Zeitschr.  f.  Hyg.,  Bd.  xxxv,  1900,  pp.  1-58, 
i  plate. 

('oi).  ZABOLOTNY.    See  vn. 

('oi).  BORDET,  J.,  ET  GENGOU,  O.  Recherches  sur  la 
coagulation  du  sang  et  les  serums  anti- 
coagulants. Ann.  de  l"Inst.  Pasteur,  T.  xv, 
1901,  pp.  129-144. 

('oi).  BORDET,  J.,  ET  GENGOU,  O.  Sur  1'existence  de 
substances  sensibilisatrices  dans  la  plupart 
des  serums  antimicrobiens.  Ann.  de  1'Inst. 
Pasteur,  1901,  T.  xv,  pp.  289-302. 

('oi).  BORDET,  J.  Sur  le  mode  d'action  des  serums 
cytolytiques  et  sur  1'unite  de  1'alexine  dans 
un  meme  serum.  Ann.  de  1'Inst.  Pasteur, 
T.  xv,  1901,  pp.  303-318. 

Coi).  GRUBER,  MAX.  Zur  Theorie  der  Antikorper. 
i.  Ueber  die  Antitoxin-Immunitat.  II. 
Ueber  Bakteriolyse  und  Haemolyse.  Miin- 
chener  med.  Wochenschr.,  Bd.  XLVIII,  1901, 
pp.  1,827-1,830,  pp.  1880-1884,  pp.  1,924-1,986. 
Not  seen. 

('02).  MARX,  E.  Die  experimentelle  Diagnostik, 
Serumtherapie  und  Prophylaxe  der  Infec- 
tionskrankheiten.  [Bibliothek  v.  Coler,  Bd. 
XL]  Berlin  (A.  Hirschwald),  1902,  pp.  vn, 
296,  mit  2  Taf. 


('04).  WASSERMANN,  A.  Immune  sera,  haemolysins, 
cytotoxins,  and  precipitins.  English  trans- 
lation by  Charles  Bolduan.  New  York,  John 
Wiley  &  Sons ;  London,  Chapman  &  Hall, 
1904,  pp.  ix,  77. 


C8o). 
C8o). 

C8i). 
C8i). 

(•81). 
C83). 

C87). 


('89) 
('93) 
('94) 
('95  ) 
('03). 


XXII.    Attenuation,  Virulence. 

PASTEUR,  Louis.  De  I'atJtenuation  du  virus  du 
cholera  des  poules.  C.  R.  des  se.  de  1'Acad. 
des  sci.,  T.  xci,  1880,  pp.  673-680. 

CHAUVEAU,  A.  Des  causes  qui  peuvent  faire 
varier  les  resultats  de  1'inoculation  char- 
bonneuse  sur  les  moutons  algeriens.  In- 
fluence des  quantites  des  agents  infectants. 
Applications  a  la  theorie  de  I'inimnnite.  C. 
R.  des  se  de  1'Acad.  des  sci.,  T.  xc,  1880,  pp. 
1,526-1,530. 

PASTEUR,  Louis.  Le  vaccin  des  oharbon.  C. 
R.  des  se.  de  1'Acad.  des  sci.,  T.  xcn,  1881, 
pp.  666-668. 

PASTEUR,  CHAMBERLAND,  ET  Roux.  De  {'at- 
tenuation des  virus  et  de  leur  retour  a  la 
virulence.  C.  R.  des  se.  de  1'Acad.  des  sci., 
T.  xcn,  1881,  pp.  429-435. 

CHAUVEAU,  A.  De  i'attenuation  des  effets  des 
inoculations  yirulentes  par  1'emploi  de^tres 
petites  quantites  de  virus.  C.  R.  des  se.  de 
1'Acad.  des  sci.,  T.  xcn,  1881,  pp.  844-848. 

CHAMBERLAND  ET  Roux.  Sur  I'attenuation  de 
la  bacteridie  charbonneuse  et  de  ses  germes 
sous  1'influence  des  substances  antiseptiques. 
C.  R.  des  se.  de  1'Acad.  des  sci.,  T.  xcvi, 
1883,  Paris,  pp.  1,410-1.412. 

METCHNIKOFF,  EuE.  Sur  I'attenuation  des 
bacteridies  charbonneuses  dans  le  sang  des 
moutons  refractaires.  Ann.  de  1'Inst.  Pas- 
teur, T.  i,  1887,  pp.  42-44. 

MACE.    See  xxm. 

D'ARSONVAL  AND  CIIARRIN.    See  XXXTI. 

ROGER.    See  xxxii. 

KLEPZOFF.    See  xxxni. 

FUHRMANN,  FRANZ.  Ueber  Vinilenzsteigerung 
eines  Stanwnes  des  Vibrio  Cholerae  asia- 
ticae.  Sitzungsber.  d.  kaiserl.  Akad.  d. 
Wissensch.  Mathematisdh-Naturwissensch. 
Klasse.,  Bd.  cxn,  Heft,  vm,  Abt.  in,  1003, 
pp.  267-284.  With  bibliography  of  15  titles. 


XXIII.    Pigments.    Green  Bacteria. 

('73).  LANKESTER,  E.  RAY.  On  a  peach-coloured 
bacterium,  Bacterium  rubescens,  n.  s. 
Quart.  Jour.  Micro.  Sci.,  1873,  vol.  xin,  n. 
s.,  pp.  408-425.  2  plates  (colored). 

('80).  VAN  TIEGHEM,  PH.  Observations  sur  des 
bacteriacees  vertes,  sur  des  phycochro- 
macees  blanches,  et  sur  les  affinites  de  ces 
deux  families.  Bull  de  la  Soc.  Bot.  de 
France,  Tome  xxvn,  1880,  pp.  174-179. 

('82).  ENCELMANN,    TH.    W.      Zur    Biologic    der 
Schizomyceten.      Bot.    Zeitung,    40    Jahrg., 
1882,  col.  321-325  and  337-34L 
Describes  a  gi-een  organism  as  Bacterium  chlorinum.    It 
is  believed  to  contain  chlorophyll. 

('87).  SCHOTTELIUS.    See  xv. 

('87).  PROVE,  OSKAR.  Micrococcus  ocliroleucus  eine 
neue  chromogene  Spaltpilzform.  Beitr.  zur 
Biol.  der  Pflanzen.,  Bd.  iv,  Hft.  3,  1887,  pp. 
409-439,  i  plate. 


PIGMENTS,    GREEN    BACTERIA. 


237 


('89).  MACK.  Sur  la  recuperation  de  la  vitalite  cles 
cultures  de  bacteries  par  passages  sur  cer- 
tains milieux.  Soc.  des  sc.  dc  Nancy  pour 
1888,  Series  n,  T.  ix,  Fasc.  xxll,  pp.  xxix- 
xxx  and  79-83,  Paris,  1889. 

The  medium  used  was  potato.  Bacteria  which  have  lost 
their  power  to  form  pigment  on  agar  or  to  infect  frogs 
will  often  regain  these  functions  if  cultivated  for  a  time 
on  potato. 

('S'O.  Zon>,  W.  Ueber  Pilz farbstoffe.  iv.  Vor- 
kommen  ei.ner  Lipochroms  bed  Spaltpilzen. 
Dot.  Zeitting,  Bd.  XLVII,  1889,  col.  89-92. 

('89).  ZOPF,  W.  Ueber  das  niikrochemischen  Ver- 
•haken  von  Fettfarbstoffen  und  Fettfarbstoff- 
Jialtigen  Organen.  Zeitschr.  f.  wissensch. 
Mikroskopie,  1889,  Bd.  vi,  pp.  172-177. 

("89).  SCHEIBENZUBER,  D.  Ein  Bacillus  mlt  brauner 
Verfarbung  der  Gelatine.  Allgemeinen 
Wiener  medicinischen  Zeitung,  Jahrg. 
xxxiv,  1889,  pp.  171-172,  Wien,  1889.  Also 
a  separate,  7  pp. 

This  organism  liquefies  gelatin.  It  was  isolated  from 
spoiled  eggs. 

('90).  BEHR,  P.  Ueber  eine  nicht  mehr  farbstoff- 
bildende  Rasse  des  Bacillus  der  'blauen 
Milch.  Centralb.  f.  Bakt.,  Bd.  vni,  1890,  pp. 
485-487- 

('90).  CLAESSEN,  HEINR.  Ueber  einen  indigoblauen 
Farbstoff  erzeugenden  Bacillus  aus  Wasser. 
Centralb.  f.  Bakt.,  Bd.  vn,  1890,  pp.  13-17. 

('91).  DANGEARD,  P.  A.  Contribution  a  1'etude  des 
Baoteriacees  vertes  (Eubacillus  gen.  nov.). 
Le  Botaniste,  Ser.  n,  1891,  fasc.  4,  pp.  151- 
160,  avec.  planche.  Also  C.  R.  des  se.  de 
1'Acad.  des  soi.,  T.  cxn,  1891,  pp.  251-253. 
Review  in  Centralb.  f.  Bakt.,  x  Bd.,  1891, 
PP.  745-747- 

Daugeard  found  his  green  spore-bearing  organism 
(Eubacillus  miiltisporus)  on  the  walls  of  a  culture  flask 
used  for  the  growth  of  fresh-water  algse,  where  it  made  a 
felt  of  very  long  slender  flexible  filaments.  He  states 
that  it  contains  chlorophyll  distributed  through  the  pro- 
toplasm. The  sporiferous  filaments  may  be  branched. 
The  spores  are  a  distinct  green.  Its  habitat  is  among 
fresh-water  algce  in  the  vicinity  of  Caen,  France. 

To  the  second  section  of  his  genus  he  would  add  the 
following  forms;  described  by  Klein :  Bacillus  de  Bary- 
anus,  B.  Solmsii,  B.  Peroniella,  B.  macrosporus,  and  B. 
limosus,  all  of  which  are  sporiferous,  the  spores  being 
blue-green.  These  spores  are  all  said  to  be  impregnated 
with  chlorophyll. 

("91).  SI<ATER,  CHARLES.  On  a  red  pigment  forming 
organism,  Bacillus  corallinus.  The  Quar- 
terly Jour,  of  Microsc.  Sci.,  1891,  vol.  32, 
n.  s.,  No.  cxxvn,  pp.  409-416,  i  plate. 

The  author  says  budding  and  branched  forms  occur  in 
liquid  media. 

('91).  BEYERINCK,  M.  W.  La  biologic  d'une  bac- 
terie  pigmentaire.  Archives  neerlandaises, 
T.  xxv,  1891,  Livr.,  314.  See  also  Die 
Lebensgeschichte  einer  Pigmentbakterie. 
Bot.  Zeitung,  1891,  Nos.  43,  45,  46,  and  47, 
columns  705-712,  741-752,  757-77O,  and  773- 
781,  with  I  plate. 

This  paper  describes  the  morphology  and  biology  of 
Bacillus  cyaneofuscus,  which  was  first  found  causing 
damage  to  the  glue  of  a  gelatin  factory  and  afterwards  in 
other  nlaces.  "  This  microbe  is  also  the  cause,  or  at  least 
one  of  the  causes,  of  a  deterioration  quite  frequent  in 
Holland  cheeses,  especially  those  known  as  '  Kdam,'  a 
change  which,  under  the  name  of  'blueing,'  is  much 
dreaded  by  makers  of  these  products." 

This  organism  belongs  to  the  chromoparous  group  of 
pigment-forming  bacteria,  i.  e.,  it  is  itself  colorless,  but 
excretes  the  pigment.  The  first  form  of  the  pigment  is  a 
blue-green  substance,  which,  under  the  microscope,  ap- 
pears as  solid-blue  spherites,  mixed  in  with  the  bacteria. 


("91).  OVERBECK,  A.  Zur  Kenntniss  der  Fettfarb- 
stoff-Produktion  foei  Spaltpilzen.  Nova 
Acta  d.  K.  Leop.  Carol.  Deutschen  Akad.  d. 
Naturf.,  Bd.  LV,  No.  7,  pp.  399-416.  i  plate. 
Halle,  1891.  Also  a  separate. 

('91).  BEIJERINCK,  M.  W.  De  Levensgeschiedenis 
eener  Pigmentbacterie.  Versl.  en  Mededeel. 
der  koninkl.  Akad.  van  We-tensch.  Afdeeling 
Naturkunde,  3  Reeks,  8  Deel,  1891,  pp. 

(3Q7)-(3I5). 
Paper  deals  with  Bacillus  cyaneofuscus. 

('91).  GESSARD,  C.  Des  races  du  bacille  pyocyanique. 
Ann.  de  1'Inst.  Pasteur,  T.  v,  1891,  pp.  65-78. 

Organism  produces  two  pigments:  (a)  Pyocyanin  (b) 
fluoresciue,  and  probably  a  third. 

("91).  ZOPF,  W.  Ueber  Ausscheidung  von  Fettfarb- 
stoffen  (Lipochromen)  seitens  gewisser 
Spaltpilze.  Ber.  d.  deutsoh.  bot.  Gesellsch., 
Bd.  ix,  pp.  22-28,  1891. 

('92).  GESSARD,  C.  Sur  la  fonction  fluorescigene  des 
microbes.  Ann.  de  I'lnst.  Pasteur.  T.  vi, 
1892,  pp.  801-823. 

('92).  CHARRIN  ET  PHYSALIX.    See  xv. 

('92).  ROHRER.  Ueber  die  Pigmentbildung  des 
Bacillus  pyocyaneus.  Centralb.  f.  Bakt.,  xi 
Bd.,  1892,  pp.  327-335. 

('92).  OKADA,  K.  Ueber  einen  rothen  Farbstoff 
erzeugenden  Bacillus  (Bacillus  rubellus) 
aus  Fussbodenstaub.  Centralb.  f.  Bakt.,  xi 
Bd.,  1892,  pp.  1-4,  mit  I  colored  Taf. 

(*93)-  ERNST,  HAROLD  C.  The  Bacillus  pyocyaneus 
pericarditidis.  The  American  Journal  of 
the  Medical  Sciences,  1893,  October,  No. 
258,  pp.  396-402.  Rev.  in  Centralb.  f.  Bakt., 
Bd.  xv,  1894,  p.  559. 

This  organism  is  motile  by  means  of  a  polar  flagellum, 
and  the  writer  has  changed  the  name  to  Bacterium  peri- 
carditidis (Ernst).  It  produces  no  pigment  soluble  in 
chloroform. 

('93)-  VOCES,  O.  Ueber  einige  im  Wasser  vorkom- 
mende  Pigmentbakterien.  Centralb.  f.  Bakt., 
Bd.  xiv,  1893,  pp.  301-315- 

An  account  of  Bacillus  cceruleus  and  other  blue  or  vio- 
let pigment-forming  organisms. 

('93).  D'ARSONVAL  AND  CHARRIN.    See  xxxn. 

('94).  BORODONI-UFKREDUZZI.     See  XLVII. 

('94).  TEISSIER,  P.  J.  Etude  des  proprietes  chromo- 
genes  permanentes  ou  facultatives  de  cer- 
tains .microbes  pathogenes  ou  saprophytes 
cultives  sur  1'albumiine  de  J'ceuf  ooagule. 
Arch,  de  med.  experim.  et  d'anatomie  path- 
ologique,  Tome  vi,  1894.  No.  2,  pp.  315-327. 

('94).  GUIGNARD  ET  SAUVAGEAU.  Sur  un  nouveau 
microbe  chromogene,  Je  Bacillus  chlorora- 
phts.  C.  R.  des  se.  et  mem.  de  la  Soc.  de 
Biol.,  Paris,  se.  x,  T.  I,  22  Dec.,  1894,  pp. 
841-843. 

('95).  SCHNEIDER,  PAUL.  Die  Bedeuitung  der  Bac- 
terienfarbstoffe  fur  die  Unterscheidung  der 
Arten.  Arbeiten  aus  dem  bakt.  Institut 
der  T«chnischen  Hoclischtile  zu  Karlsruhe. 
Bd.  i,  Hft.  2,  1895,  pp.  201-232.  i  fig.  and 
i  Tafel. 

Conclusions  :  (i)  Distinguishable  in  part  by  diverse  be- 
havior in  solvents.  (2)  The  same  organism,  under  like 
conditions,  always  produces  the  same  pigment.  (3)  Two 
sorts  may  produce  the  same  pigment.  (4)  Most  of  those 
sorts  which  produce  apparently  the  same  pigment,  and 
are  much  alike  in  other  ways,  may  be  distinguished 
easily  by  the  different  reactions  of  the  pigments. 

('95).  DANGEARD,  P.  A.  Observations  sur  le  groupc 
des  bacteries  vertes.  Ann.  de  micrograpihie, 
1895,  T.  VIT,  pp.  67-69. 

Considers  Bact.  viride,  van  Tieghem,  to  belong  to  the 
algie. 


238 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('95).  THUMM,  K.  Beitrage  zur  Biologic  der  fluor- 
escirenden  Bakterien.  Arbeiten  aus  dem 
Bak't.  Inst.  der  Tec'hnischen  Hoohschule  zu 
Carlsruhe,  Bd.  i,  1895,  pp.  291-377. 

The  following  summary  of  the  most  important  results 
is  introduced  in  extenso,  owing  to  the  difficulty  of  pro- 
curing the  original  paper : 

1.  All  fluorescent  bacteria  show  in  alkaline  gelatin, 
first  a  sky-blue,  later  a  moss-green  fluorescence,  and  with 
the  latter  a  yellowing  of  the  substratum.    Old  cultures, 
with  the  exception  of  those  of  Bacillus  fluorescens  puti- 
dus,  are  orange-red,  with  a  dark -green  fluorescence. 

2.  All  these  colors  are  due  to  one  yellow  pigment,  a 
concentrated  watery  solution  of  which  is  orange  yellow, 
a  dilute  one  yellow.     Both  fluids  are  blue  fluorescent,  but, 
upon  the  addition  of  an  alkali,  become,  according  to  the 
concentration,  dark-green  or  moss-green  fluorescent. 

3.  All  species  produce  the  same  pigment. 

4.  All  are  alkali  formers.     The  production  of  ammonia 
in  many  species  is  considerable,  and  it  is  due  to  the  pres- 
ence of  this  alkali  that  the  blue  fluorescence  gives  place 
to  the  green. 

5.  The  view  of  Naegeli,  I^edderhose,  and  Kunz  of  the 
formation  of  a  leuco-pigment,  and  their  attempt  to  trace 
back   the  different  colorings   to    oxidation  phenomena, 
has  not  been  confirmed. 

6.  Bacillus  pyocyaneus  Ernst,  contrary  to  the  view  of 
the  other  investigators,  forms  only  one  pigment. 

7.  Ill  potato  cultures  and  on  acid  gelatin,  the  same  col- 
oring matter  is   produced  as  in   alkaline  media.     The 
green  fluorescence  is,  in  every  case,  caused  by  the  action 
of  the  ammonia. 

8.  The  a.  aud£  forms  of  Bacillus  pyocyaueus  differ  only 
by  the  amount  of  ammonia  produced  not  by  a  difference 
in  the  pigment  termed.     The  a  form  is  a  good,  the  j3  form 
a  poor  alkali  producer.    When  ammonia  is  added  to  a  cul- 
ture of  the  ft  form,  it  resembles  a  culture  of  the  a  form. 

9.  When  an  acid  producer  and  a  fluorescent  form  are 
grown  together  in  the  same  culture,  the  yellow  pigment 
appears  normally,  but  there  is  no  fluorescence. 

10.  All  species  have  the  power  of  oxidizing  grape  sugar 
with  the  production  of  an  acid.    The  ammonia  formed 
later  neutralizes  this. 

11.  The  addition   of  sodium  formate  to   the  ordinary 
nutrient  gelatin  causes  an  increased  ammonia  produc- 
tion, 

12.  In  hydrochinon  gelatin  all  species  produce  a  brown- 
ish-red color,  due  to  the  action  of  ammonia  on  the  hydro- 
chinon.    This  may,  under  certain  circumstances,  be  used 
as  a  test  for  ammonia. 

13.  The  behavior  of  the  different  species  in  media  con- 
taining different  organic  substances  is  so  characteristic 
that  it  may  serve  as  a  valuable  means  of  differentiating 
related    species.      Ammonium  succinate,  or  asparagin, 
affords  good  nourishment  for  all  species. 

14.  According  to  the  source  of  the  carbon  or  nitrogen, 
the  same  organism  is  an  abundant  or  scanty  alkali  pro- 
ducer; e.g.,  Bacillus  pyocyaneus  Ernst  is  a  poor  alkali 
former  in  nutrient  gelatin,  but  a  good  one  in  ammonium 
succinate. 

15.  Each  organism  shows  manifestations  of  life   only 
when  it  comes  in  contact  with   the  oxygen   of  the  air. 
Hence,  only  there  do  we  find  pigment  and  formation  of 
ammonia. 

16.  For  the  formation  of  the  pigment,  calcium  chloride 
is  entirely  unessential,  but  magnesium  sulfate  and  potas- 
itim  phosphate  are  of  the  greatest  importance.    Gessard's 
view,  that  only  phosphoric  acid  is  absolutely  necessary, 
is  erroneous,  nor  may  it  ever  be  concluded  that  absence  of 
fluorescence  it  due  to  absence  of  phosfates. 

17.  The  substitution   of  calcium  for  magnesium,  and 
vice  versa,  does  not    influence  the  development  of  the 
different  species,  but  in  the  formation  of  the  pigment, 
calcium  can  not  take  the  place  of  magnesium. 

18.  The  blue  color  of  a  fluid  culture  of  Bacillus  pyocy- 
aneus, in  the  absence  of  phosphoric  acid,  is  nevercaused 
by  pyocyanin,  as  Gessard  assumed,  but  is  referable  only 
to  refraction  phenomena. 

19.  The  least  amount  of  potassium  phosfate  or  mag- 
nesium sulfate  is  sufficient  for  the  formation  of  the  fluor- 
escent   pigment.     Where,    in    such    nutrient   solutions, 
mostly  blue  fluorescence  is  to  be  observed,  there  has  been 
a  decreased  amount  of  ammonia  produced  because  of  the 
small  supply  of  nutrient  salts  in  the  solution. 

20.  Bacterium  syncyaneum  has  the  power  of  forming 
two  pigments,  a  fluorescent   and  a  steel-blue  one.     The 
former  agrees  with  that  of  the  other  fluorescent  species. 
The  latter  varies  from  steel-blue  to  brownish  black,  ac- 
cording to  the  reaction  of  the  medium. 

21.  The  fluorescence  of  the  £  form  of  Bacterium  syu- 
cyaneum  may  be  produced  by  cultivating  it  in  ammonium 
lactate  and  transferring  to  nutrient  gelatin. 


('96).  BIEL,  WILHELM.     Ueber  einen  schwarzes  Pig- 
nientbildenden    Kartoffelbacillus.      Centralb. 
f.  Bakt.,  2  Abt.,  Bd.  n,  1896,  pp.  137-140. 
('96).  SCHEURLEN.     Geschichtliche   und   experiment- 
elle  Studien  tiber  den  Prodigiosus.    Archiv. 
f.  Hyg.,  1896,  Bd.  xxvi,  pp.  1-31. 
('96).  GORINI,    C.      Ueber    die    schwarzen    pigment - 
ibildenden   Bakterien.     Centralb.   f.   Bakt.,   i 
Abt.,  Bd.  xx,  1896,  p.  94. 

('97).  EWART,  A.  J.     Bacteria  with  asshnilatory  pig- 
ments,   found    in    the    tropics.      Annals    of 
Botany,  vol.  xi,  1897,  pp.  486-487. 
Author  found  seven  greenish  bacteria  in  water  at  Buit- 
enzorg,  Java,  showing  a  faint  evolution  of  oxygen  when 
exposed  to  light.    These  were  B.  chlorinum  and  Strepto- 
coccus varians,  two  forms  closely  resembling  van  Tieg- 
heni's  B.  virens  and  Bact.  viride;  another  large  bacillus, 
somewhat  resembling  van  Tieghem's  B.  vireus,  and  two 
Spirilla.     The  red   Bacterium  photometricum,  which  is 
common  in  Java,  gives,  on  treatment  with  hot  alcohol 
and  extraction  with  benzine,  a  green  dye  which  seems  to 
be  identical  with  chlorophyll. 

('97).  THIRY,  G.  Contribution  a  1'etude  du  poly- 
•chromisme  bacterien.  Bacille  et  Cladothrix 
polychromes;  cristaux  colores.  Arch,  de 
physiol.,  1897,  No.  2,  pp.  284-288. 

('97).  NEUMANN,  RUDOLF.  Studien  iiber  die  Varia- 
ibilitat  der  Farbstofifbildung  bei  Mikrococcus 
pyogenes  a  aureus  ( Staphylococcus  pyo- 
genes  aureus)  und  einigen  anderen  Spalt- 
pilzen.  Arch,  f.  Hyg.,  Bd.  xxx,  1897,  pp. 
1-31.  i  table. 

"  Die  eiue  Race   kann  also  aus  der  anderen   entstehen 
und  in  eine  andere  iibergefiihrt  werdeu." 


('98). 


('98). 
('98). 

('99). 


('99). 


('99). 


WARD,  H.  MARSHALL.  A  violet  bacillus  from 
the  River  Thames.  Annals  of  Botany,  vol  xn, 
1898,  pp.  59-74.  One  double  plate  in  color. 

NIEDERKORN.     See  xv. 

RUZICKA.    See  xv. 

JORDAN,  EDWIN  O.  The  production  of  fluores- 
cent pigment  by  bacteria.  Botanical  Gazette, 
vol.  xxvn,  pp.  19-36,  1899. 

JORDAN,  E.  O.  Bacillus  pyocyaneus  and  its 
pigments.  Jour.  Exper.  Med.,  vol.  iv,  Nos. 
5  and  6,  1899,  pp.  627-647.  ' 

BOLAND,  G.  W.    Ueber  Pyocyanin  den  Wauen 
Farbstoff  des  Bacillus  pyocyaneus.    Centralb. 
1  Bakt.,  xxv  Bd.,  1899,  pp.  897-902,  with  i 
curve. 
Very  probably  B.  pyocyaneus  forms  two  pigments. 

('oo).  THIRY,  GEORGES.     Bacille  polychrome  et  Acti- 
nomyces  mordore.     Recherches   biologiques 
sur  lee  bacteries  bleues  et  violettes.     Poly- 
chromisme.      Corps    bacteriens    €t    cristaux 
colores.        Matiere     colorante     cristallisee. 
Travaux  du  lab.  dMiyg.  et  de  1'inst.  serothe- 
rapique  de  1'Univer.  de  Nancy.     Paris,  J.  B. 
Balliere  et  fils,  1900,  pp.  vm,  154,  7  plates. 
Contains  also  a  bibliography  of  141  titles  on  pigment- 
forming  bacteria. 

('oo).  KUNTZE,  W.  Ein  Beitrag  zur  Kenntnis  der 
Bedingungen  der  Farbstoffbildung  des 
Bacillus  prodigiosus.  Zeitschr.  f.  Hyg.,  Bd. 
xxxiv,  Hft.  i,  1900,  pp.  169-184.  Rev.  in  Cen- 
tralb. f.  Bakt,  xxvin  Bd.,  1900,  pp.  602-604. 

With  a  solution  made  up  of  100  parts  pure  water,  i  to  2 
asparagin,  2  to  4  c.  p  grape  sugar,  and  0.2  dipotassium 
phosphate,  the  author  obtained  a  fairly  good  growth  of 
B.  prodigiosus  without  color.  With  the  same  solution 
and  a  grape  sugar  not  quite  pure,  there  was  always  a 
formation  of  pigment.  The  white  bacteria  became  pig- 
mented  in  a  few  hours  on  potato,  or  on  adding  a  trace  of 
(o.ooi )  of  MgSO4.  This  substance  contaminated  the  sugar 
first  used. 


PIGMENTS;    REDUCTION    AND   OXIDATION;    NITRIFICATION,    ETC.         239 


Coo).  KRAUSE.    See  xv. 

Coo).  CHAMOT,  E.  M.,  AND  THIRY,  G.     Studies  on 

chromogenic   'bacteria.      I.      Notes    on    the 

pigment  of  Bacillus  polychromogenes.     Bot. 

Gaz.,  vol.  xxx,   1900,  pp.  378-393.     16  figs. 

Also  a  separate. 
COT).  GESSARD,   C.     Variete   melanogene   du   bacille 

pyocyanique.     Ann.   de   1'Inst.    Pasteur,   T. 

xv,  1901,  pp.  817-831. 
('02).  GESSARD,  C.     Essai  sur  la  biologic  du  bacille 

pyocyanique.     Ann.  Inst.  Pasteur,  Paris,  T. 

xvi,  1902,  pp.  313-330. 
('02).  PETROW,   N.    Uber   einen   neuen   roten   Farb- 

stoff-bildenden    Bacillus.    Arb.    a.    d.    Bact. 

Institut  der  techn.  Hochschule  zu  Karlsruhe, 

ii  Bd.,  3  Heft,  1902,  pp.  271-291,  with  i  plate. 

Describes  Bacillus  suhkilieusis. 

('02).  LoEw,  O.,  AND  KOZAI,  Y.  Ueber  Ernahrungs- 
verhaltnisse  beim  Bacillus  prodigiosus.  Bull, 
of  the  College  of  Agric.,  Tokyo  Imperial 
Univ.,  vol.  v,  1902,  No.  2,  pp.  137-141.  Also 
a  separate. 

A  favorable  medium  for  production  of  pigment  and 
bacteriolytic  enzyme  is  composed  of  peptone  i  per  cent, 
sodium  acetate  0.2  per  cent,  and  asparagin  0.2  per  cent  in 
water. 

('03).  MARSH.    See  vi. 

('03).  PAPENHAUSEN,  HUBERT.    Uber  die  Bedingun- 

gen  der  Farbstoffbildung  bei  den  Bakterien. 

Arb.  a.  d.  Bact.  Inst.  der  tech.  Hochschule 

zu  Karlsruhe,  in  Bd.,  i  Heft,  1903,  pp.  43-79. 

Bibliog.  of  20  titles. 

Twenty-two  species  experimented  upon.  Oxygen  is 
very  necessary  for  the  production  of  the  pigments.  The 
other  conditions  for  optimum  production  of  pigment  vary 
greatly  in  different  species. 

('04).  HEi'FERAN,  MARY.  A  comparative  and  ex- 
perimental study  of  bacilli  producing  red 
pigment.  A  dissertation  submitted  to  the 
faculties  of  the  Graduate  Schools  of  Arts, 
Literature  and  Science,  in  candidacy  for  the 
degree  of  Doctor  of  Philosophy.  The  Uni- 
versity of  Chicago.  Printed  in  Jena  by 
Gustav  Fischer,  1904,  pp.  55.  Bibliog.  of  77 
tides. 

('04).  LEONARD,  ETHEL  L.  Bacterium  cyaneum:  A 
new  chromogenic  organism.  The  Johns 
Hopkins  Hospital  Bulletin,  vol.  xv,  1904, 
PP.  398-400. 


XXIV.     Reduction  and  Oxidation. 

('87).  SPINA,    A.      Bacteriologische    Versuche    mit 

gefanbten     Nahrsubstanzen.       Centralb.     f. 

Bakt.,  ii  Bd.,  Jena,  1887,  Nos.  2-3,  pp.  71-75. 

Reduction  processes  of  bacteria  in  presence  of  methv- 

len  blue,  etc. 

('87).  CAHEN,  FRITZ.  Ueber  das  Reduktionsver- 
mogen  der  Bakterien.  Zeitschr.  f.  Hyg., 
Bd.  ii,  1887,  pp.  386-396. 

('9l).    WlNOGRADSKY.      See  XXV. 

('94).  FERMI,  CLAUDIO,  AND  MONTESANO,  GIUSEPPE. 
Ueber  die  Dekomposition  des  Amygdaliirs 
durch  Mikroorganismen.  Centralb.  f.  Bakt., 
Bd.  xv,  1894,  pp.  722-727. 

('96).  SMITH,  THEOBALD.  Reduktionserscfaeinungen 
bei  Bakterien  und  ihre  Beziehungen  zur 
Hakterienzelle,  nebst  Bemerkungen  tiber  Re- 
duktionserschcinungen  in  steriler  Bouillon. 
Centralb.  f.  Bakt,  xix  Bd.,  1896,  pp.  181-187. 


('99)-  MUELLER,  FRIEDR.  Ueber  reduzierende  Eigen- 
schaften  von  Bakterien.  Centralb.  f.  Bakt., 
xxvi  Bd.,  1899,  pp.  51-63. 

The  pigment  used  must  be  soluble  in  water,  and  must 
not  poison  the  bacteria.  Author  uses  methylene  blue 
and  litmus.  Under  literature,  n  papers  are  cited. 

('99).  ROTHBERGER.     See  xvni. 

('99).  MUELLER,  FRIEDRICH.  Ueber  das  Reduktions- 
vennogen  der  Bakterien.  Centralb.  f.  Bakt., 
xxvi  Bd.,  1899,  pp.  801-819. 

Author  used  methyleue  blue,  litmus,  indigo-carmiu  and 
rosanilin  acetate  in  various  media  with  many  organisms. 

Coi).  GRAN.    See  xxv. 

('02).  CATHCART,  EDUARD,  UND  HAHN,  MARTIN. 
Ueber  die  reduzierenden  Wirkungen  der 
Bakterien.  Arch.  Hyg.,  Munchen,  Bd.  XLFV, 
1902,  pp.  295-321. 

('02).  EMMERLING,  OSCAR.  Die  Zersetztmg  stick- 
stofffreier  organischer  Substanzen  durch 
Bakterien.  Braunschweig  (F.  Vieweg  & 
S.),  1902,  pp.  ix,  141,  mit  7  Taf. 

Not  seen. 

('03).  VAN  DELDEN,  A.  Beitrag  zur  Kenntniss  der 
Sulfatreduction  durch  Bacterien.  Centralb. 
f.  Bakt.,  2  Abt.,  xi  Bd.,  1903,  pp.  81-94  and 
113-119,  i  heliotype  pi. 


XXV.    Nitrifying  and  Denitrifying  Organisms, 
Use  of  Free  Nitrogen. 

('66).  WORONIN.  Ueber  die  bed  der  Sohwarzerle  und 
der  gewohnlichen  Lupine  auftretenden 
Wurzelanschwellungen.  Memoires  de 
1'Acad.  imp.  St.  Petersb.,  7  Serie,  T.  x,  1866. 
See  also  Ann.  des  sci.  nat.  Bot.,  5  se.,  T.  vn 
pp.  73-86,  i  plate. 

('77)-  SCHLOESING,  TH.,  ET  MUENTZ,  A.  Sur  la 
nitrification  par  les  ferments  organises.  C. 
R.  des  se.  de  1'Acad.  des  sci.,  Paris,  1877, 
T.  LXXXIV,  p.  301 ;  and  T.  LXXXV,  p.  1,018. 

('79).  SCHLOESING,  TH.,  ET  MUENTZ,  A.  Recherches 
sur  la  nitrification.  C.  R.  des  se.  de  1'Acad. 
des  sci.,  Paris,  T.  LXXXIX,  1879,  pp.  891-894 
and  1,074-1,077. 

('82).  DEH£RAN  ET  MAQUENNE.  Sur  la  reduction 
des  nitrates  dans  la  terre  arable.  C.  R.  des 
se.  de  1'Acad.  des  sci.,  1882,  T.  xcv,  pp.  691- 
693,  732-734,  854-856. 

('82).  DUPETIT,  G.,  ET  GAYON,  U.  Sur  la  fermenta- 
tion des  nitrates.  C.  R.  des  se.  de  1'Acad. 
des  sci.,  T.  xcv,  pp.  644-646,  Paris,  1882. 

('82).  DUPETIT,  G.,  ET  GAYON,  U.  Sur  la  transforma- 
tion des  nitrates  en  nitrites.  C.  R.  des  se. 
de  1'Acad.  des  sci.,  T.  xcv,  Paris,  1882,  pp. 
1,365-1,367. 

('83).  DUPETIT,  G.,  ET  GAYON,  U.  Quelques-unes  des 
conditions  les  plus  favorables  a  la  fermenta- 
tion des  nitrates.  Mem.  de  la  soc.  des  sci. 
physiques  et  naturelles  de  Bordeaux,  2  se., 
T.  v,  1883,  Extr.  Proces-verbaux  se.  du  23 
nov.,  1882,  pp.  ni-iv. 

('83).  SPRINGER,  A.  Reduction  of  nitrates  by  fer- 
ments. Am.  Chem.  Jour.,  vol.  iv,  pp.  452- 
453,  1883. 

('86).  GAYON  ET  DUPETIT.    Recherches  sur  la  reduc- 
tion des  nitrates  par  les  innniments  petits., 
1886.     Paris,?  Publisher? 
Not  seen. 


240 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('86).  MUNRO,  J.  H.  M.  The  formation  and  destruc- 
tion of  nitrates  and  nitrites  in  artificial  solu- 
tions, and  in  river  and  well  waters.  Jour. 
Chem.  Soc.  Trans.,  London,  1886,  vol.  xi.ix, 
pp.  632-681. 

('88).  BEYERINCK.  Die  Bakterien  der  Papiliona- 
ceenknollohen.  Bot.  Zeitung,  Jahrg.  XLVI, 
1888,  col.  725-735,  741-750,  757-771,  781-790, 
797-804,  2  plates.  Review  in  Centralb.  f. 
Bakt.,  Bd.  v,  1889,  pp.  804-805. 

('88).  HELLRIECEL,  H.,  UND  WIU-'ARTH,  H.  Unter- 
suchungen  iiber  die  Stickstoffnahrung  der 
Graimineen  und  Legurninosen,  1888.  Boila- 
geheft  zu  der  Zeitsohr.  d.  Vereins  f.  d. 
Rubenzucker-Industrie  d.  Deutschen  Reichs. 
Berlin,  pp.  234,  with  6  plates. 

('90).  FRANKLAND,  PERCY  F.  The  nitrifying  process 
and  its  specific  ferment,  Ft.  i.  Phil.  Trans. 
Roy.  Soc.,  Lond.,  vol.  181  B.,  1890,  pp.  107- 
128. 

('90).  WINOGRADSKY,  S.  Recherches  sur  les  organ- 
ismes  de  la  nitrification.  Ann.  de  1'Inst. 
Pasteur,  T.  iv,  1890,  pp.  213-231,  257-275, 
and  760-771. 

Nitroinonas  can  live  on  purely  inorganic  matters, 
although  destitute  of  chlorophyll.  His  "  nitratfermeut " 
consists  of  pear-shaped  rods,  which  do  not  exceed  %  ^  in 
length,  and  are  l%  to  2  times  less  in  breadth.  The 
"  nitritorganismus "  is  4  to  5  times  larger  than  the 
"  nitratbildner,"  and  has  a  roundish  form. 

('90).  LAURENT,  EM.  Experiences  sur  la  reduction 
des  nitrates  par  les  vegetaux.  Ann.  de  1'Inst. 
Pasteur,  T.  iv,  pp.  722-744,  1800. 

('91).  GILT  AY,  E.,  UND  ABERSON,  J.  H.  Recherches 
sur  un  mode  de  denitrifieation  et  sur  le 
schizomycete  qui  la  produit.  Archives  neer- 
landaises  des  sci.  ex.  et  nat.,  1891,  T.  xxv, 
pp.  34i-36i,  i  plate. 

('9l).    SCHLOESING,    TH.,    FILS,    ET    LAURENT,    EMILE. 

Sur  la  fixation  de  1'azote  litre  par  les 
plantes.  C.  R.  des  se.  de  1'Acad.  des  soi. 
Paris,  1891,  T.  cxin,  pp.  776-778  and  1,059- 
1,060,  1892,  T.  cxv,  pp.  659-661  and  732-735. 
('91).  WINOGRADSKY,  S.  Recherches  sur  les  organ- 
ismes  de  la  .nitrification.  Ann.  de  1'Inst. 
Pasteur,  T.  v,  1891,  pp.  92-100  and  577-616. 

Winogradsky's  fourth  and  fifth  memoirs  on  nitrifying 
bacteria.  The  fourth  deals  with  the  question  of  solid 
media  suited  to  their  culture,  and  describes  the  making 
of  silicate  jelly.  The  fifth  deals  with  the  formation  and 
the  oxidation  of  nitrites  in  the  nitrification. 

('91).  BEIJERINCK,  M.  W.  Over  pphooping  van  at- 
mospherische  stikstof  in  culturen  van 
Bacillus  radicicola.  Versl.  en  Mededeel.  der 
Koninkl.  Akad.  van  Wetensch.  zu  Amster- 
dam, Afd.  Natuurkunde,  in,  8,  1891,  pp. 
(460) -(475),  i  text  fig.  Abstract  in  Koch's 
Jahresb.,  Ill  Bd.,  p.  205. 

('92).  WINOGRADSKY,  S.  Contributions  a  ;la  mor- 
phologic des  organismes  de  la  nitrification. 
Arch.  d.  sci.  biol.  publiees  par  1'Institut  Im- 
perial de  med.  exper.  a  St.  Petersbourg, 
Tome  i,  1892,  p.  108.  Reviewed  in  Ann.  de 
1'Inst.  Pasteur,  T.  vi,  1892,  pp.  458-462. 

('92).    SCHLOKSING,    TH.,    FILS,    ET    LAURENT,    EMII.E. 

.Recherches  sur  la  fixation  de  1'azote  libre 
par  'les  plantes.  Ann.  de  1'Inst.  Pasteur, 
1892,  T.  vi,  pp.  65-115. 

('92).  BERTHELOT.  Nouvelle  recherches  sur  la  fixa- 
tion de  1'azote  par  les  microbes.  C.  R.  des  se. 
de  1'Acad.  des  sci.,  1892,  T.  cxv,  pp.  569-574. 


('93).  BERTHEI.OT.       Recherches     nouvelles     sur     les 

microorganismes  fixateurs  de  1'azote.     C.  R. 

des  se.  de  1'Acad.   des   sci.,   Paris,   1893,  T. 

cxvi,  pp.  842-849. 
('94).  DUCLAUX,  E.     Sur  la  fixation   de  1'azote   at- 

mospherique.  Revue  critique.    Ann.  de  1'Inst. 

Pasteur,  1894,  T.  vm,  pp.  728-736. 
('94).  WINOGRADSKY,  S.    Sur  1'assimilation  de  1'azote 

gazeux   de   i'atmosphere   par   les    microbes. 

C.  R.  des  se.  de  1'Acad.  des  sci.,  Paris,  T. 

cxvi,  1893,  p.  1,385,  and  T.  cxvni,  1894,  Feb. 

12,  p.  353.     Rev.  in  Centralb.   f.   Bakt.,  Bd. 

xvi,  1894,  pp.  129-130. 
('95).  WINOGRADSKY,  S.     Recherches  sur  i'assimila- 

tion  de  1'azote  libre  de  ['atmosphere  par  les 

•microbes.     Arch,  des  sci.  biol.  pub.  par  1'In- 

stitut.   imperial    de   medeoine   experimentale 

a  St.   Petersbourg,  Tome   in,  No.  4,   1895, 

PP.  297-352. 
A  very  interesting  and  valuable  paper. 

('96).  WINOGRADSKY,  S.  Zur  Mikrobiologie  des 
nitrifikationsprozesses.  Centralb.  f.  Bakt., 
2  Abt.,  Bd.  ii,  1896,  pp.  415-428. 

('97).  MAZE.  Fixation  de  1'azote  libre  par  le  bacille 
des  nodosites  des  legumineuses.  Ann.  de 
1'Inst.  Pasteur,  T.  xi,  1897,  pp.  44-54. 

The  second  and  third  memoirs  are  continued  in  the 
Annales  for  1898,  under  the  title,  "  lyes  microbes  des 
nodosites  des  legumineuses." 

('98).  KONVALEWSKI,  S.  Beitrage  zur  Frage  iiber  die 
Assimilierung  von  freiem  Stickstoff  seitens 
der  Bakterien.  Russ.  Arch.  f.  Pathol.,  Bd. 
vr,  1898,  p.  251.  Rev.  in  Centralb.  f.  Bakt., 
xxv  Bd.,  1899,  pp.  771-772. 

('98).  MAZE.  Les  microbes  des  nodosites  des  legum- 
ineuses. Ann.  de  1'Inst.  Pasteur,  T.  XH, 
1898.  Second  memoir,  pp.  1-25,  i  fig.  Third 
memoir,  pp.  128-155,  2  plates. 

The  second  memoir  deals  with  the  physiology  of  the 
organism.  The  third  memoir  deals  with  the  morphology 
of  the  root-tubercle  organism.  Maz£  sa3's  it  is  patho- 
genic for  certain  animals.  In  the  animal  body  it  is  said 
to  become  nearly  round,  to  be  much  reduced  in  diame- 
ter, and  to  have  a  tendency  to  grow  in  chains. 

('99).  OMEUANSKY,  V.  Sur  la  nitrification  de  1'azote 
organique.  Arch,  des  sci.  biol.  publiees  par 
1'inst.  imp.  de  med.  exper.  a  St.  Potersbourg, 
T.  vii,  1899,  pp.  272-290. 

('99).  OMEUANSKY,  V.  Sur  la  culture  des  microbes 
nitrificateurs  du  sol.  Arch,  des  sci.  biol.  pub- 
liees par  1'inst.  imp.  de  med.  exper.  a  St. 
Petersbourg,  T.  vn,  1899,  pp.  291-302. 

(99).  WINOGRADSKY  u.  OMEUANSKY.  Ueber  den 
Einflus  der  organiisphen  Substanzen  auf  die 
Arbeit  der  nitrificierenden  Mikrobien. 
Centralb.  f.  Bakt.,  2  Abt.,  Bd.  vi,  1899,  p.  329. 

('99).  WINOGRADSKY,  S.,  ET  OMEUANSKY,  V.  L'in- 
fkience  des  substances  organiques  stir  le 
travail  des  microbes  nitrificateurs.  Arch, 
des  sci.  biol.  publiees  par  1'inst.  imp.  de  med. 
exper.  a  St.  Petersbourg,  T.  vn,  1899,  pp. 

233-271- 

('oo).  KRUEGER,  W.,  AND  SCHNEIDEWIND,  W.  Ursache 
und  Bedeutung  der  Salpeter-Zersetzung  ini 
Boden.  Landwirtschaftliche  Jahrbiicher. 
Ztschr.  f.  wiss.  Landwirtschaft,  Berlin, 

1900,  Bd  xxix,  pp.  747-770,  2  plates. 

COT).  BKUERTNCK,  M.  W.  Ueber  oligonitrophile 
Mikroben.  Centralb.  f.  Bakt.,  etc.,  Bd.  vn, 

1901,  pp.  561-582,  with  i  heliotype  plate. 

Describes  Azotobacter. 


NITRIFICATION,    ETC.;    USE   OF    FREE   CO2 ;    LUMINOSITY. 


241 


('01).  GRAN,  II.  H.  Studien  uber  Meeresbakterien. 
i.  Reduction  von  Nitraten  und  Nitriten. 
Bcrgens  Museums  Aarbog.,  1901,  No.  10, 
PP.  23. 

Three  new  species  are  described  :  liacillus  repens,  B. 
trivialis,  B.  Heusenii. 

('or).  MAASSEN,  ALBERT.  Die  Zerzetzung  der  Nitrate 
und  der  Nitrite  durch  die  Bakterien.  Ein 
Beitrag  zum  Kreislatif  des  Stickstoffs  in  der 
Natur.  Arbeit,  a.  d.  K.  Gesundheitsamt., 
Bd.  xvin,  1901,  pp.  21-77. 

('02).  MOORE,   GEO.   T.     Bacteria  and   the   Nitrogen 
Problem.     Yearbook  Dept.  Agric.,  1902,  pp. 
333-342,  6  plates.     Also  a  separate. 
Pure  cultures  of  the    root-tubercle  organism  are  ab- 
sorbed on  cotton,  which  isthen  dried,  wrapped  in  tin  foil, 
and  thus  distributed  to  the  agriculturist.     He  is  directed 
to  throw   the  cotton  into  a   pail  of   water,  to  which  is 
added  small  packages  of  nutrient  salts.     After  48  hours 
the  fluid  is  filled  with  the  bacterial  growth.     The  legum- 
inous seeds  are  then  drenched  with  it,  and  sown  after  dry- 
ing in  the  shade. 

('02).  SMITH,  R.  GREIG.  Notes  on  Vibrio  denitrifi- 
•oans,  Sewerin.  Proc.  Linnean  Soc.,  New 
South  Wales,  for  the  year  1901.  Sydney, 
1902,  vol.  xxvi,  Pt.  i,  pp.  118-121.  i  plate. 

('02).  OM£I,IANSKI,  W.  Kleinere  Mitteilungen  uber 
Nitrifikationsmikroben.  I.  Die  Kultur  des 
Nitritbildners  auf  Papierscheiben.  n.  Wird 
schweflige  und  phosphorige  Satire  durch 
Nitrobacter  oxydiert?  in.  Scheiden  die 
Nitritmikroben  eine  Oxydase  aus?  Cen- 
tralb. Bakt,  Abt.  2,  Bd.  ym,  1902,  pp.  785- 
787;  Bd.  ix,  1902,  pp.  63-65,  pp.  113-117,  mit 
i  Taf. 

('02).  BEIJERINCK,  M.  W.,  UND  VAN  DELDEN,  A. 
Ueber  die  Assimilation  des  freien  Stick- 
stoffs durch  Bacterien.  Centralb.  Bakt., 
Abt.  2,  Bd.  ix,  1902,  pp.  3-43. 

("02).  BUHLERT,  H.  Untersuchungen  uber  die  Art- 
einheit  der  Knollchenbakterien  der  Legumi- 
noscn  und  iiber  die  landwirtschaftliche  Be- 
deutung  dieser  Frage.  Habilitationsschr. 
Halle  (Druck  v.  Wischan  u.  Wettengel), 
1902,  p.  55.  Centralb.  Bakt.,  Abt.  2,  Bd.  ix, 
1902,  pp.  148-153,  226-240,  and  273-285. 

("02).  Iln.TNER,  L.  Ueber  die  Impfung  der  Legumin- 
osen  mit  Reinkulturen.  D.  landw.  Presse, 
Berlin,  Bd.  xxix,  1902,  pp.  119-120. 

('02).  NOBBE,  FRIEDRICH,  UND  RICHTER,  L.  Ueber 
den  Einfluss  des  Nitratstickstoffs  und  der 
Humussubstanzen  auf  den  Impfungserfolg 
bei  Leguminosen.  Landw.  Versuchsrat., 
Berlin,  Bd.  LVI,  1902,  pp.  441-448. 

("03).  BERSTEYN,  P.  Uber  einige  in  den  Kulturen 
zur  Reinziichtung  der  Nitratbildner  regel- 
massig  auftretende  Bakterienarten.  Arb.  a.  d. 
Bact.  Inst.  der  tech.  Hochschule  zu  Karls- 
ruhe, in  Bd.,  i  Heft,  1903,  pp.  81-100. 

The  following  new  species  are  described  :  Bacterium 
comes,  B.  modestum,  B.  debile  and  Ps.  humicola.  None 
of  these  species  will  grow  in  media  entirely  free  from 
organic  matter. 

('03).  BENECKE,  W.,  UND  KEUTNER,  J.  Ueber  stick- 
stoffbindende  Bakterien  aus  der  Ostsee. 
Ber.  d.  deutsch.  bot.  Gesellsoh.,  Bd.  xxi, 
Heft  6,  pp.  333-346,  Berlin,  1903,  4  text  figs. 

('05).  MOORE,  GEORGE  T.  Soil  Inoculation  for  Le- 
gumes. Bureau  of  Plant  Industry,  U.  iS. 
Department  of  Agriculture,  Bull.  71,  Jan.  23, 
1005,  pp.  72,  pi.  9. 


XXVI.     Use  of  Free  Carbon  Dioxide. 

("87).  HUEPPE,  FERDINAND.  Ueber  Chlorophyllwirk- 
ung  cli'lorophyllfreier  Pflanzen.  Tageblatt 
der  60  versammlung  deutscher  Naturforscher 
u.  Aerzte  in  Wiesbaden,  1887,  pp.  244-245. 

('99).    WlNOGRAUSKY  U.  OM^LIANSKY.      See  XXV. 

('02).  NATHANSOHN.    See  u. 

('03).  BEIJERINCK,  M.  W.,  UND  VAN  DEUDEN,  A. 
Ueber  eine  farblose  Bakterie,  deren  Kohlen- 
stoffnahrung  aus  der  atmospharischen  Luft 
herruhrt.  Centralb.  f.  Bakt.,  2  Abt.,  Bd.  x, 
No.  2,  1903,  pp.  33-47.  See  also  Versl.  Wis. 
Nat.  Afd.  K.  Acad.  Wet.,  Bd.  xi,  1903,  pp. 
450-465  (Dutch)  ;  and  Proc.  Sci.  K.  Acad. 
Wet.,  Bd.  v,  1903,  pp.  398-413  (English), 
Amsterdam. 

"We  will  use  the  name  Bacillus  oligocarbophilus  to 
designate  a  colorless  bacterium  whose  carbon  needs,  in 
the  dark  as  well  as  in  the  light,  are  satisfied  by  a  not  yet 
exactly  known  carbon  compound,  or  compounds,  of  the 
air,  out  of  which  compound  this  organism  must  also  cre- 
ate the  necessary  energy  for  its  life  processes."  It  is 
stated  that  this  substance  is  not  carbon  dioxide. 

('04).  BEIJERINCK,  M.  W.  Ueber  die  Bakterien, 
welche  sioh  im  Dunkeln  mit  Kohlensaure 
als  Kohlenstoffquelle  ernahren  konnen. 
Centralb.  f.  Bakt.,  2  Abt.,  xi  Bd.,  1904,  No. 
20-22,  pp.  593-599- 


XXVII.    Luminous  Bacteria. 


('75). 


PFLUEGER,  E.  Beitrage  zur  Lehre  von  der 
Respiration.  Sec.  5.  Die  Phosphorescenz 
der  lebendigen  Organistnen  und  ihre  Bedeu- 
tung  fur  die  Principien  der  Respiration. 
Pfliiger's  Archiv,  1875,  Bd.  x,  pp.  275-300. 

("80).  LASSAR,  O.  Die  Mkrococcen  der  Phosphores- 
cenz. Pfluger's  Arch.  f.  die  gesarn.  Physio!., 
1880,  Bd.  xxi,  pp.  104-108. 

('85).  NUEECH.     Ueber  leuohtende  Bacterien.     Bale, 

1885. 
Not  seen. 

('87).  FISCHER.  Bakteriologische  Untersuchungen 
auf  einer  Reise  nach  Westindien.  u.  Ueber 
einen  lichtentwickelnden,  im  Meerwasser 
gefundenen  Spaltpilz.  Zeitschr.  f  .Hyg.  Bd. 
n,  1887,  pp.  54-92,  and  Nachtrag,  pp.  92-95. 

('87).  FORSTER,  J.  Ueber  einige  Eigensohaften  leuch- 
•bender  Bakterien.  Centralb.  f.  Bakt.,  Bd. 
it,  1887,  pp.  337-340. 

('87).  KATZ,  OSCAR.  Preliminary  remarks  on  phos- 
phorescent bacteria  from  sea-water.  Proc. 
Linn.  Soc.,  New  South  Wales,  vol.  11,  Pt. 
n,  1887,  PP- 331-336. 

('87).  DUCLAUX.  Sur  les  microbes  phosphorescents. 
Revue  critique.  Ann.  de  1'Inst.  Pasteur,  T.  i, 

1887,  pp.  489-492. 

('88).  DUBOIS,  R.  Sur  le  role  de  la  symbiose  chez 
certains  animaux  marins  lumineux.  C.  R. 
des  se.  de  1'Acad.  des  sci.,  T.  evil,  Paris, 

1888,  pp.  502-504. 

('88).  FISCHER,  B.  Ueber  einen  neuen  lichtentwick- 
elnden Bacillus.  Centralb.  f.  Bakt.,  1888, 
in  Bd.,  pp.  105-108,  and  pp.  137-141. 

('89).  BEYERINCK,  M.  W.  Le  photobacterium  lumi- 
nosum,  bacterie  lumineuse  de  la  mer  du 
nord.  Arch.  neer.  des  sci.  ex.  et  nat.,  T. 
xxin,  1889,  pp.  401-415. 


242 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('89).  BEYERINCK,  M.  W.  Les  bacteries  lumineuses 
dans  leurs  raports  avec  1'oxygene.  Arch, 
neer.  des  sci.  ex.  et  nat,  T.  xxm,  1889,  pp. 
416-427. 

('89).  GIARD,  A.,  UND  BILLET,  A.  Observations  sur 
la  tnaladie  phosphorescente  des  Talitres  et 
autres  Crustaces.  C.  R.  de  la  Soc.  de  biol., 

1889,  Tome  i,  pp.  593-597- 

('89).  LEHMANN,  K.  B.  Studien  fiber  Bacterium 
phosphorescens  Fischer.  Centralb.  f.  Bakt., 
v  Bd.,  1889,  pp.  785-79L 

("90).  GIARD,  A.  Nouvelles  reoherohes  sur  les  bac- 
teries lumineuses  pathogenes.  C.  R.  de  la 
Soc.  de  biol.,  1890,  Tome  u,  pp.  188-191. 

('90).  BEIJERINCK,  M.  W.  Over  lichtypedsel  en 
plastisch  voedsel  van  lichtbacterien.  Versl. 
en  Meded.  d.  Kon.  Akad.  v.  Wetenschappen, 
Aft.  Natuurk.  Der.de  Reeks,  Deel  vn,  1890, 
pp.  239-302.  I  text  figure. 

('90).  BILLET,  A.  Contribution  a  1'etude  de  la  mor- 
phologic et  du  developpement  des  bac- 
teriacees.  Bull,  scientifique  de  la  France  et 
de  la  Belgique,  public  par  Giard,  T.  xxi, 

1890,  pp.    1-289,   plates  9.     Bibliography  of 
662  titles.    Also  a  separate. 

The  bacterium  of  In  in  incus  sand  fleas  is  described  in  a 
uote  on  p.  144  as  Bacterium  Giardi  Billet. 

('91).  BEYERINCK,  M.  W.  Sur  1'aliment  photogene 
et  1'aliment  plastique  des  bacteries  lumi- 
neuses. Arch,  neerl.  des  sci.  ex.  et  nat.,  T. 
xxiv,  1891,  pp.  369-442. 

('91).  KATZ,  OSCAR.  Zur  Kenntniss  der  Leuchtbak- 
terien.  Centralb.  f.  Bakt.,  1891,  ix  Bd.,  pp. 
I57-I63,  199-204,  229-234,  258-264,  311-316, 
343-349- 

('92).  RUSSELL,  H.  L.    Impfungsyersuche  mit  Giard's 
pathogenem     Leuchtbacillus.       Centralb.     f. 
Bakt.,  xi  Bd.,  1892,  pp.  557-559- 
Author  obtained  no  conclusive  evidence  as  to  the  patho- 
genic nature  of  this  organism. 

('92).  EIJKMANN,  C.  Lichtgevende  Bacterien. 
GeneeskundigTijdschriftvoorNederlandsch- 
Indie.  Deel  xxxn,  Aflevering  4,  Batavia  en 
Noordwijk,  1892,  pp.  109-115.  Also  a  sepa- 
rate. Rev.  in  Centralb.  f.  Bakt.,  xn  Bd., 
1892,  pp.  656-657. 

('93).  DUBOIS,  RAPHAEL.  Extinction  de  la  lumi- 
nosite  du  Photobacterium  sarcophilum  par 
la  lumiere.  C.  R.  des  se.  et  mem.  de  la  soc. 
de  Biologic,  Paris,  1893,  9  ser.,  T.  v,  pp.  160- 
161. 

This  organism  was  isolated  from  the  luminous  flesh  of 
a  dead  rabbit.  It  was  cultivable  in  a  synthetic  medium 
made  as  follows  :  Ordinary  water,  loo  ;  asparagiue,  i  ; 
glycerin,  i ;  potassium  phosphate,  o.io ;  sea  salt,  3.  After 
some  months  in  this  medium  the  luminosity  diminished. 
Kxposed  to  the  light  for  some  days,  at  about  10°  C.,the 
culture  takes  on  a  fine  orange  yellow  color,  becomes 
opaque,  and  loses  its  luminosity  except  at  the  edges. 
Transfers  may  be  made  from  this  yellow  growth  readily, 
but  the  resulting  cultures  are  not  luminous.  In  the  dark, 
however,  such  cultures  return  after  some  days  to  their 
orginal  transparent  color,  and  again  become  luminous. 

('95).  KUTSCHER.  Zur  Phosphorescens  der  Elbvib- 
rionen.  Centralb.  f.  Bakt.,  xvm  Bd.,  1895, 
p.  424. 

('98).  FRANKLAND,  PERCY.  The  action  of  bacteria  on 
the  photographic  plate.  Centralb.  f.  Bakt., 
xxiv  Bd.,  1898,  pp.  609-612. 

Only  the  luminous  sorts  are  able  to  act  through  glass. 
The  action  of  non  luminous  species  is  probably  due  "to 
the  evolution  of  volatile  chemical  substances."  The  lat- 
ter exposures  were  long — several  days. 


('99).  HENNEBERG,  W.     Leuchtbakterien  als  Krank- 

heitserreger  bei  Schwammucken.     Centralb. 

f.  Bakt.,  xxv  Bd.,  1899,  pp.  649-650. 
('99).  BARNARD,  J.  E.     Photogenic  Bacteria.    Trans. 

of  the  Jenner   Inst,   London,   1899,   second 

series,  pp.  81-112.    2  plates. 

"  Spectroscopically  the  light  emitted  by  the  photogenic 
organisms  examined  by  me  is  confined  to  a  very  small 
portion  of  the  visible  spectrum  ;  never  extending  into 
the  ultra-violet  or  infra-red.  Visually  it  only  includes 
the  green  and  blue,  and  photographically  it  extends  very 
slightly  further  into  the  violet.  The  economic  value  of 
such  a  light  source  is  obvious,  and  it  is  a  matter  of  great 
practical  importance  to  determine  its  method  of  produc- 
tion." 

The  writer  experimented  with  13  species. 

('02).  McKENNEY,  R.  E.  B.  Observations  on  the 
conditions  of  light  production  in  luminous 
bacteria.  Proc.  of  Biol.  Soc.  of  Wash., 
Nov.  20,  1902,  vol.  xv,  pp.  213-234.  Also  a 
•separate.  Bibliog.  of  35  titles. 

('02).  BARNARD,  J.  E.,  AND  MACFADYEN,  ALLAN.  On 
Luminous  Bacteria.  Annals  of  Botany,  vol. 
xvi,  Dec.,  1902,  pp.  587-588. 

("03).  MOLISCH,  HANS.  Vienna  Acad.  Sci.,  1903. 
Luminous  bacteria  for  Safety  Lamp. 
Science,  1903,  p.  719. 

Paper  read,  but  not  yet  printed.  Note  in  Science  made 
from  a  Renter's  telegram. 

('04).   GORIIAM,  F.  P. 

See  a  preliminary  note  in  Ceutralb.  f.  Bakt.,  2Abt., 
XIII  Bd. 


XXVIII.     Hydrogen  Sulphide  and  Otherwise 
Unclassified  By-Products. 

('79).  MIQUEL,  P.  De  la  fermentation  sulfhydriquc. 
Bull,  de  la  Soc.  ohim.,  T.  xxxn,  1879,  P- 
127-131. 

('89).  HOLSCHEWNIKOFF.  Uaber  die  Bildung  von 
Schwefelwasserstoff  dtirch  Bakterien. 
Fortschr.  d.  Med.,  1889,  Bd.  vii,  pp.  201-213. 

('91).  NENCKI,  M.  Die  isomeren  Milohsauren  als 
Erkennungsmittel  oinzelner  Spaltpilzarten. 
Centralb.  f.  Bakt,  Bd.  ix,  1891,  pp.  304-306. 

('gi-'93).  RAY-PAILHADE.  Reoherohes  experiment- 
ales  sur  le  philothion.  Paris,  1891.  Le 
phiilothion  et  le  soufre.  Assoc.  frang.  pour 
1'av.  des  sci.,  Congres  de  Besangon,  1893. 
Part  i,  pp.  193,  250,  and  302. 

('92).  SOMMARUGA,  E.  Ueber  Stoffwechselproducte 
von  Mikroorganismen.  Zeitschr.  f.  Hyg., 
Bd.  xii,  1892,  pp.  273-297. 

('92).  PETRI,  R.  J.,  u.  MAASSEN,  A.  Ueber  die  Bil- 
dung von  Schwefelwasserstoff  durch  die 
Krankheitserregenden  Bakterien  imter  be- 
sonderer  Berucksichtigung  des  Schweine- 
rothlaufs.  Veroffent.  d.  k.  Gesundheitsamtes, 
xvi  Jahrg.,  1892,  p.  119. 

('93).  STAGNITTA-BALISTRERI.  Die  Verbreitung  der 
Schwefelwasserstoffbildung  unter  den  Bak- 
terien. Arch,  fur  Hyg.,  Bd.  xvi,  1893,  pp. 
10-34- 

For  demonstration  of  sulphuretted  hydrogen  in  gela- 
tin, the  author  adds  iron  saccharate,  tartrate,  or  acetate 
to  the  gelatin.  For  fluids  he  uses  lead  acetate  paper. 

('93).  RUBNER.     Ueber    den    Modus    der    Schwefel- 

wasserstoffbildttng  bei  Bakterien.     Arch.   f. 

Hyg.,  Bd.  xvi,  1893,  pp.  53-72. 
('93).  RUBNER.      Die    Wanderungen    des    Schwefels 

im    Stoffwechsel    der    Bakterien.      Arch.    f. 

Hyg.,  1893,  Bd.  xvi,  pp.  78-100. 


HYDROGEN    SULPHIDE,    ETC.;   ACTION    OF    LIGHT   ON    BACTERIA. 


243 


('93)-  PETRI,  R.  J.,  UND  MAASSEN,  ALBERT.  Beitrage 
zur  Biologic  der  kranlcheitserregenden  Bak- 
tcricn  ins  besondcre  iiber  die  Bildung  von 
Schwefelwasserstoff  dtirch  dieselben  unter 
vornehmlicher  Berucksichtigung  des  Schwei- 
nerothlaufs.  Arbeiten  aus  dem  Kaiserl. 
Gesundlieitsamte,  Bd.  vm,  1893,  pp.  318-356. 

M:my  parasitic  ami  saprophytic  forms  have  been  tested. 
All   produce  sulphuretted  hydrogen  when  cultivated  in 


cua    |iwuwc   viuiwumwu    ujru&wgvzi    wucu    \.uiuvtucu    in 

presence  of  suitable  sulphur  compounds,  i.  e. ,  such  as  are 
loosely  bound.  The  reduction  of  litmus  and  indigo  in 
solid  cultures  is  ascribed  to  this  gas.  It  is  believed  that 
all  bacteria  are  capable  of  producing  it,  and  that  the  dis- 
tinction into  producers  and  non-producers  of  sulphur- 
etted hydrogen  must  be  abandoned. 

('93).  PETRI,  R.  J.,  UND  MAASSEN,  ALBERT.  Wekere 
Beitrage  zur  Schwefelwasserstoffbildung 
aerober  Bakterien  und  kurze  Angaben  uber 
Merkaptanbildung  derselben.  Arbeken  aus 
dem  Kaiserlichen  Gesundheitsamte,  Bd.  vin, 
1893,  pp.  490-506. 

('93).  RUBNER,  M.  Ueber  das  Vorkommen  von  Mer- 
captan.  Arohiv.  f.  Hyg.,  1893,  Bd.  xix,  pp. 
136-193. 

('94).  WENZELL,  W.  T.  A  contribution  to  the 
knowledge  of  bacteriologic  chemistry.  Jour. 
Am.  Med.  Asso.,  1894,  vol.  xxm,  pp.  901- 
003. 

('96).  DUCLAUX,  E.  Stir  les  odeurs  de  putrefaction. 
Revue  critique  Ann.  de  1'Inst.  Pasteur,  T. 
x,  1896,  pp.  59  to  64. 

('97).  MORRIS,  MAX.  Sttidien  fiber  die  Produktion 
von  Schwefelwasserstoff,  Indol  und  Merkap- 
tan  bei  Bakterien.  Aroh.  f.  Hyg.,  xxx  Bd., 
1897,  PP.  304-3". 

Mercaptan  was  demonstrated  by  use  of  *' isatinschwe- 
felsiiure,"  which  is  colored  green  by  this  substance. 

('02).  BANNING,  FRIEDRICH.  Zur  Kenntniss  der  Oxal- 
saurebildung  durch  Bakterien.  Centralb.  f. 
Bakt.,  Abt.  2,  Bd.  vm,  1902,  pp.  520-525,  pp. 
556-567,  mit  i  Taf. 


XXIX.     Action  of  Light  on  Bacteria. 

('77).  DOWNES  AND  BLUNT.    Researches  on  the  ef- 
fect of  light  upon  bacteria  and  other  organ- 
isms.    Proc.  Roy.  Soc.,  vol.  xxvi,  No.  184, 
1877,  6  Dec.,  pp.  488-500.    London. 
Downes  and  Blunt  determined  germicidal  action  to  be 
associated  chiefly  with  the  actinicraysof  the  spectrum. 

('78).  DOWNES  AND  BLUNT.  On  the  influence  of 
light  upon  protoplasm.  Proc.  Roy.  Soc.,  vol. 
xxvni,  No.  191,  1878,  19  Dec.,  pp.  199-212. 
('78).  TYNDALL,  J.  Note  on  the  influence  exercised 
by  light  on  organic  infusions.  Proc.  Roy. 
Soc.,  London,  vol.  xxvni,  pp.  212-217.  Na- 
ture, vol.  xix,  1879,  p.  210. 

Negative  results.    Used  flasks,  and    organisms   grew 
after  exposure  to  sunlight. 

('81).  TYNDALL,  JOHN.  On  the  arrestation  of  in- 
fusorial life  by  solar  light.  Br.  asso.  for  the 
advancement  of  sci.  Rep.,  1881,  pp.  450-451. 
Nature,  vol.  xxiv,  p.  466,  1881. 

Tyndall's  results   were  due    to    the  way  in  which   he 
experimented,  i.  e.,  with./fa*fc  cultures. 

('82).  ENGELMANN,  TH.  W.  Bacterium  photome- 
tricum.  Ein  Beitrag  zur  vergleichenden 
physiologie  des  Licht-  und  Farbensinnes. 
Onderzoekingen  gedaan  in  het  Physiologisch 
Laboratorium  der  Utrechtsche  Hoogeschool, 
Derde  Reeks,  vn,  Aflev.  n,  1882,  pp.  252- 
290,  I  plate. 


('84).  DOWNES,  A.,  AND  BLUNT,  T.  P.    The  influence 
of  light  on  bacteria.    Trans.  Roy.  Soc.,  Vic- 
toria, vol.  xx,  pp.  1-2,  1884. 
Reply  to  Jamieson. 

('85).  DUCLAUX,  E.    Influence  de  la  lutniere  du  soleil 

sur  la  vitalite  des  germes  de  microbes.     C. 

R.   des  se.   de  1'Acad.   des  sci.  T.   C.,   1885, 

pp.  119-121. 
('85).  DUCLAUX,  E.    Influence  de  la  lumiere  du  soleil 

sur  la  vitalite  des  micrococcus.     C.  R.  des 

se.  de  1'Acad.  des  sci.,  Paris,  T.  ci,  1885,  pp. 

395-397- 
('86).  ARLOING,  S.     Influence  de  la  lumiere  blanche 

et  de  ses  rayons  constituants  sur  le  devei- 

oppement  et  les  proprietes  du  Bacillus  an- 

thracis.    Arch,  de  physiol.  normale  et  pathol., 

1886,  3  sen,  T.  vn,  pp.  209-235. 
('86).  STRAUS,  I.     Note   sur  1'action   de  la  lumiere 

solaire  sur  les  spores  du  Bacillus  anthracis. 

C.  R.  des  se.  et  mem.  de  la  soc.  de  Biologic. 

Paris,  8  se.,  T.  ill,  1886,  pp.  473-474. 
('86).  DOWNES,  ARTHUR.     On  the  action  of  sunlight 

on   Micro-organisms  with   a   demonstration 

of  the  influence  of  diffused  light. — Proceed- 
ings of  the  R.   Society,  London,   1886,  vol. 

XL,  pp.  14-22. 
('87).  DUCLAUX,  E.     Action   de   la  lumiere  sur  les 

microbes.     Ann.  de  1'Inst.  Pasteur,  1887,  T. 

i,  pp.  88-92. 
('87).  Roux,  E.    De  1'action  de  la  lumiere  et  de  1'air 

sur  les  spores  de  la  bacteridie  du  charbon. 

Ann.  de  1'Inst.  Pasteur,  T.  i,  1887,  pp.  445- 

452. 
('88).  ENGELMANN,   TH.   W.     Die    Purpurbakterien 

und    ihre    Beziehungen    zurn    Liohte.    Bot. 

Zeitung,  1888,  Jahrgang  XLVI,  col.  661,  677, 

693,  and  709. 
('88).  ENGELMANN,  TH.  W.    Ueber  Bacteriopurpurin 

und       seine       physiologische       Bedeutunjj. 

Pfliiger's  Archiv.,  1888,  Bd.  XLII,  pp.  183-186. 
('89).  RAUM,  JOHANNES.     Der  gegenwartige   Stand 

unserer   Kenntnisse   iiber   den   Einfluss  des 

Lichtes   auf   Bacterien   und   auf   den   thier- 

ischen  Organismus.     Zeit.   f.   Hyg.,   Bd.  vi, 

1889,  pp.  312-368. 
This  paper  contains  a  bibliography  of  7  pages. 

('89).  PANSINI,  S.  Dell'azione  della  luce  solare  sui 
microorganismi.  Reyista  d'igiene  practica 
e  sperimentale,  Napoli,  1889,  pp.  69-101.  Re- 
view in  Ann.  de  Micr.,  1890,  p.  516. 

('90).  SAVERIO.     L'influenza   della   temperatura   sull" 
azione    microbicida   della    luce.     Ann.    dell' 
Inst.  d'ig.  di  Roma. 
Not  seen. 

('90).  JANOWSKI,   TH.     Zur    Biologic    der   Typlius- 
bacillen.     Centralb.   f.   Bakt.,  vm  Bd.,   1890, 
pp.  167-172,  193-199,  230-234,  and  262-266. 
Discusses  the  action  of  sunlight. 

('91).  GEISLER,  F.  K.    On  the  action  of  light  on  bac- 
teria.    (Russian.)     Wratsch,   1891,   No.  36, 
PP.  793-797 
Not  seen. 

('92).  MOMONT.    See  xxxv. 

('92).  KOTLJAR,  E.    Zur  Frage  iiber  den  Einfluss  des 

Lichtes  auf  Bakterien.    Wratsch,  1892,  Nos. 

39  and  40.     Rev.  in  Centralb.   f.  Bakt.,  xn 

Bd.,   1892,  p.  836.     Also  in  Ann.   de  1'Inst. 

Pasteur,  T.  vn,  1893,  p.  430. 


244 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('92).  CHMELEWSKY.  Zur  Frage  fiber  die  Wirkung 
des  Sonnen-  und  elektrischen  Lichtes  auf 
die  Eiterbakterien.  Wratsch,  1892,  No.  20. 
Reviewed  in  Centralb.  f.  Bakt.,  Bd.  xn,  pp. 
174-175,  1892. 

('92).  GEISLER,  THEODOR.  Zur  Frage  fiber  die  Wir- 
kung des  Lichtes  auf  Bakterien.  Centralb. 
f.  Bakt,  xi  Bd.,  1892,  pp.  161-173. 
('92).  BUCHNER,  H.  Ueber  den  Einfluss  des  Lichtes 
auf  Bakterien.  Centralb.  f.  Bakt.,  xi  Bd., 
1892,  pp.  781-783- 

Deals  with  question  of  effect  of  light  on  bacteria  sus- 
pended in  water.  "  The  result  of  all  these  experiments 
points  to  the  conclusion  that  light  exerts  a  powerful  dis- 
infecting influence  upon  the  named  bacterial  sorts  when 
these  are  suspended  in  water."  The  experiments  were 
made  on  typhoid  bacilli,  B.  coli  communis,  B.  pyocyan- 
eus,  cholera  vibrios,  and  various  bacteria  of  decay. 

('92).  BUCHNER,  H.    Ueber  den  Einfluss  des  Lichtes 
auf  Bakterien.    11  Mitth.    Centralb.  f.  Bakt, 
Bd.  xn,  1892,  Heft  7-8,  pp.  217-219,  I  fig. 
Author  obtained  bacterial  letters  and  figures  on  agar 
and  gelatin  plates  by  covering  a  part  and  exposing  to 
sunshine.    Light   passed  through  water   still  possesses 
active  bactericidal  powers. 

('93).  BUCHNER,  H.  Ueber  den  Einfluss  des  Lichtes 
auf  Bakterien  und  fiber  die  Selbstreinigung 
der  Flfisse.  Arch.  f.  Hyg.,  Bd.  xvn,  1893, 
pp.  179-204. 

('93).  RICHARDSON,  ARTHUR.  The  action  of  light  in 
preventing  putrefactive  decomposition ;  and 
in  the  formation  of  hydrogen  peroxide  in 
organic  liquids.  Jour.  Chem.  Soc.,  Trans- 
actions, London,  1893,  vol.  LXIII,  pp.  1,109- 
1,130. 

D'ARCY,  R.  F.,  AND  HARDY,  W.  B.  Note  on 
the  oxidizing  powers  of  different  regions  of 
the  spectrum  in  relation  to  the  bactericidal 
action  of  light  and  air.  Jour,  of  Physio!., 
i894-'95,  vol.  xvn,  pp.  3QO-393- 
WARD,  H.  MARSHALL.  The  Action  of  light  on 
bacteria.  Phil.  Trans.  Royal  Soc.,  Lond., 
vol.  185  (1894),  pp.  961-986.  Also  a  sepa- 
rate. 4to.,  25  pp.,  i  plate.  See  also  Revue 
Sci.,  1894. 

('94).  FISCHER,  BERNHARD.  Die  Bakterien  des 
Meeres  nach  den  Untersuchungen  der  Plank- 
tonexpedition  unter  gleichzeitiger  Beriick- 
sichtigung  einiger  alterer  und  neuerer  Un- 
tersuchungen. Centra*,  f.  Bakt.,  Bd.  xv, 
1894,  pp.  657-666. 

Shows  that  sunlight  tends  to  reduce  number  of  bacteria 
in  upper  layers  of  the  sea.  There  are  always  more  bac- 
teria in  upper  layers  of  the  sea  at  sunrise  than  in  the 
afternoon.  There  are  also  in  the  daytime  a  great  many 
more  nt  a  depth  of  10  meters  than  near  the  surface.  The 
depth  to  which  the  bactericidal  action  of  the  sunlight 
penetrates  depends  on  its  intensity,  duration  of  action, 
clearness  of  the  water,  etc.  It  probably  reaches  down 
several  meters.  Cultures  of  various  water  bacteria  were 
killed  in  a  short  time  when  exposed  to  the  midday  sun  in 
August,  the  sun's  rays  being  first  passed  through  one-half 
meter  of  sea  water. 

('94).  DIEUDONNE,   A.     Beitrage  zur  Beurteilung  der 
Einwinkung     des     Lichtes     auf     Bakterien. 
Anbeiten  a.  d.  Kaiserl.  Gesundheitsarnte,  Bd. 
ix,  1894,  Heft  2,  pp.  405-413.     Rev.  in  Cen- 
tralb. f.  Bakt.,  xvii  Bd.,  1895,  pp.  646-647. 
The  red  end  of  the  spectrum  (between  the  lines  D  and 
R)  has  no  injurious  effect.     In  the  green   (between  lines 
K  and  F)  there  is  a  distinct  inhibiting  action.     In  the 
blue-violet  and  ultra-violet  the  bactericidal  action  is  most 
marked.     The  action  of  the  light  is  directly  on  the  bac- 
teria.    Bacteria  inoculated  in  plates  already  exposed  to 
the  light  grew  just  as  well  as  in  the  control  plates.    Light , 
which  has  had   the   heat    rays    removed    by    filtration 
through  alum  solution,  possesses  the  same  germicidal 
action. 


('94).  DIEUDONNE,  A.  Ueber  die  Bedeutung  des 
Wasserstoffsuperoxyds  fur  die  bakterien- 
todtende  Kraft  des  Lichtes.  Arbeiten  aus 
dem  kais.  Gesundheits-Anit,  Bd.  ix,  1894, 
PP.  537-540. 

Ascribes  bactericidal  effect  of  light  in  great  part  to  the 
formation  of  hydrogen  peroxide  in  the  culture-medium. 

('94).  ENGELMANN.     See  xix. 

('94).  D'ARSONVAL  AND  CHARRIN.     See  xxxin. 

('96).  BECK,  M.,  u.  SCHULTZ,  P.  Ueber  die  Ein- 
wirkung  sogen.  monochromatischen  Lichtes 
auf  die  Bacterienentwicklung.  Zeitschr.  f. 
Hyg.,  Bd.  xxin,  1896,  pp.  490-496. 

('99).  KEDZIOR,  LAURENZ.  Ueber  den  Einfluss  des 
Sonnenliohtes  auf  Bakterien.  Arch.  f.  Hyg., 
Bd.  xxxvi,  1899,  pp.  323-334.  Rev.  in  Cen- 
tralb. f.  Bakt,  Bd.  xxvn,  1900,  pp.  203  and 
759- 

Sunlight  also  destroys  in  an  atmosphere  of  hydrogen, 
although  less  actively. 

('oi).  SIMONCINI,  G.  B.,  E  VIOLA,  D.  L'influenze 
deiU'innaffiamento  sul  contenuto  batterico 
delle  polveri  di  strada.  Ann.  d'igicne  sper., 
Roma,  vol.  xi,  1001,  pp.  373-392. 

Bibliog.  of  21  titles.  The  bactericidal  action  of  liglit 
was  greater  on  the  moistened  dust  of  the  street  than  on 
the  dry  dust. 


XXX.    Effect  of  Electricity. 


('91) 


('91) 


('93) 


SPILKER,  W.,  UND  GOTTSTEIN,  A.  Ueber  die 
Vernichtung  von  Mikroorganismen  durch 
die  Induktionselektrickat.  Centralb.  f. 
Bakt,  ix  Bd.,  1891,  pp.  77-88. 

FERMI,  CLAUDIO.  Ueber  die  Reinigung  der 
Abwasser  durch  Elektriciitat  Arch,  fur 
Hyg.,  Bd.  xm,  1891,  pp.  207-228. 

KRUEGER,  S.  Ueber  den  Einfluss  des  constan- 
ten  elektrisohen  Stromes  auf  Wachsthum 
der  Bakterien.  Zeitschr.  f.  klin.  med.,  Bd. 
xxn,  1893,  pp.  191-207. 

Krueger's  conclusions  are  :  The  inhibition  or  destruc- 
tion of  the  bacteria  is  due  to  the  liberation  of  ions  in  the 
fluid. 

('93).  BURCI,  E.,  E  FRASCANI,  V.  Contribute  allo 
studio  dell'azione  battericida  della  corrente 
continua.  —  Atti  della  Soc.  Tosc.  di  Scienze 
nat  Pisa.  Mem.,  vol.  xn,  1893.  pp.  99-119. 

('94).  D'ARSONVAL  AND  CHARRIN.     See  xxxni. 

('96).  FRTEDENTHAL,  H.  Ueber  den  Einfluss  des 
elektrischen  Stromes  auf  Bakterien.  Krit- 
isches  Referat  Centralb.  f.  Bakt.,  xix  Bd., 
1896,  pp.  319-324- 

('96).  GOTTSTEIN,  A.  Ueber  den  Einfluss  des  elek- 
trischen Stromes  auf  Baktcrden.  Centralb. 
f.  Bakt.,  xix  Bd.,  1896,  pp.  602-605. 

('96).  FRIEDENTHAL,  H.  Ueber  den  Einfluss  des 
Induktionselektrizitat  auf  Bakterien.  Krit- 
isches  Referat  Centralb.  f.  Bakt,  xx  Bd., 
1896,  pp.  505-508. 

('96).  MARMIER,  L.  A.  Les  toxines  et  relectricite. 
Ann.  de  1'Inst  Pasteur,  T.  x,  1896,  pp.  469- 
480. 

('99).  THIELE,  HERMANN,  UND  Wou>,  KURT.     Ueber 
die    Einwirkung    des    elektrischen    Stromes 
auf  Bakterien.     Centralb.  f.  Bakt.,  xxv  Bd., 
1899,  pp.  650-655,  with  i  fig. 
Results  all  negative. 

('oo).  KRAUSE.    See  xv. 


ELECTRICITY;  ROENTGEN  RAYS,  ETC.;  HIGH  PRESSURE. 


245 


('01).  STREHEI.,  I  IKUMANN.  Untersuchungcn  iiber  die 
luktericide  Wirkung  des  Hochspannungs- 
funkenliohtes  nebst  Angabe  einer  Methode 
zur  besseren  Ausntitzung  der  baktericiden 
Kraft  des  Voltabogenlichtes.  D.  med. 
Wochensclir.,  Berlin,  Bd.  xxvn,  1901,  pp. 
69-72,  pp.  87-89. 

('oi).  UI.T.MANN,  JOHANNES.  Ueber  die  Einwirkung 
elektrischcn  Bogenlichts  auf  Mikroorgan- 
iMiien  in  Gegenwart  von  fluoreszierenden 
Stoffen.  Diss.  Munch  en  (Druck  v.  M. 
Ernst),  1901,  p.  17. 


XXXI.    Action  on  Bacteria  of  Roentgen  Rays,  Ra- 
dium Rays,  Etc. 

('96).  WITTLIN,  J.  Les  rayons  Rontgen  exercent-ils 
une  action  quclconque  sur  les  bacteries? 
Ann.  de  -micro.,  T.  vm,  1896,  pp.  514-515. 

Author  fiuds  that  the  Roentgen  rays  have  no  effect  upon 
bacteria. 

('96).  MINCK,  P.  Zur  Frage  iiber  die  Einwirkung 
der  Rontgen'sehen  Strahle.n  auf  Baktericn 
und  ihre  eventuelle  therapeutische  Verwend- 
barkeit.  Munchener  mediz.  Woohenschrift. 
1806,  Bd.  XLIII,  pp.  101-102  and  p.  202. 

Author  obtained  only  negative  results. 

('97).  POTT,  FRANCIS.     Concerning  .the  action  of  X- 
rays  on  cultivation  of  tubercle  Bacillus.   The 
Lancet,    London,    vol.    n,    for    1897    (55th 
year),  pp.  1,314-1,315. 
The  tubercle  bacillus  was  not  affected  by  X-rays. 

('97).  BLAISE  ET  SAMBUC.     De  Faction  des  rayons  X 
sur   Je    Pyocyaneus    et    la    bacteridie    char- 
bonneuse.     C.  R.  des  .se.  et  mem.  de  la  soc. 
de  biol.,  T.  iv,  ice  serie,  1897,  pp.  689-692. 
Little  or  no  effect  on  these  organisms. 

('97).  BEAUREGARD  ET  GUICHARD.  Action  des  rayons 
X  sur  certains  characteres  biologique  des 
microbes.  C.  R.  des  se.  et  mem.  de  la  soc. 
de  biol.,  T.  iv,  loe  serie,  1897,  pp.  803-804. 

The  bacteria  are  much  less  sensitive  than  higher  or- 
ganisms. 

('98).  RiEDER,  HERMANN.  Wirkung  der  Rontgen- 
strablen  auf  Bakterien.  Munch,  med. 
Wochensclir.,  45  Jahrg.,  1898,  pp.  101-104,  2 
text  figures  (exposed  agar  plates). 

Contrary  to  the  statements  of  various  other  experi- 
menters, this  writer  says  that  he  obtained  positive  germi- 
cidal  results  on  seven  pathogenic  organisms  by  exposures 
lasting  from  45  minutes  to  i  hour.  The  earlier  literature 
is  cited.  VolCohm's  apparatus  was  used.  The  photo- 
graphs show  the  center  of  the  agar  Petri-dish  cultures 
cleared  of  bacterial  colonies. 

('98).  WOLFENDEN,  MORRIS,  AND  FORBES-ROSS,  F.  W. 
A  preliminary  note  on  the  action  of  the 
Roentgen  rays  upon  the  growth  and  activity 
of  bacteria  and  micro-organisms.  The 
Lancet,  London,  June  25,  1898,  pp.  1,752- 
1,753- 

Bacillus  prodigiostis  011  potato  was  exposed  to  the  rays 
for  one  hour  on  several  occasions.  Growth  was  much 
greater  than  in  the  control  tubes,  and  more  pigment  was 
formed. 


('98).  RIEDER,  H.  Weitere  Mittheilung  iiber  die 
Wirkung  der  Rontgenstrahlen  auf  Bacterien 
sowie  auf  die  menschliche  Haut.  Munch, 
med.  Wochenschr.,  45  Jahrg.,  1898,  pp.  773- 
774- 

foi).  CASPARI,  W.  Ueber  die  bacterienschadigende 
Wirkung  der  Becquerelstrahlen.  Nadi  in 
Gemeinschaft  mil  Priv.  Doc.  Dr.  Aschkinass 
ausgefuhrten  Versuchen.  Arch.  ital.  biol., 
Turin,  T.  xxxvi,  1901,  p.  130. 

('02).  RIEDER,  HERMANN.  Nochmals  die  bakterien- 
todtende  Wirkung  der  Rontgenstrahlen. 
Munchener  med.  Wochenschr.,  Bd.  XLIX, 
1902,  pp.  402-406. 

('04).  PRESCOTT,  S.  C.  The  effect  of  radium  rays  on 
the  colon  bacillus,  the  diphtheria  bacillus 
and  yeast.  Science,  n.  s.,  vol.  xx,  Aug.  19, 
1904,  pp.  246-248. 

"  Radium  rays  have  no  effect  upon  fresh  cultures  of  B. 
coli,  B.  diphtheria,  or  Saccharomyces  cerevisiae  at  a  dis- 
tance of  one  centimeter  where  the  time  of  exposure  is 
less  than  90  minutes. 


XXXII.     Effect  of  High  Pressure  on  Bacteria. 

('75)-  BERT,  P.  Influence  de  1'air  comprime  sur  les 
fermentations.  C.  R.  des  se.  de  1'Acad.  des 
sci.,  Paris,  1875.  T.  LXXX,  pp.  1,579-1,582. 

('77).  BERT,  P.  De  1'emploi  de  1'oxygene  a  haute 
tension  comme  precede  d'investigation  physi- 
ologique ;  des  venins  et  des  virus.  C.  R.  des 
se.  de  1'Acad.  des  sci.,  Paris,  1877,  T.  LXXXIV, 
pp.  1,130-1,133. 

('91).  SCHAFFER  ET  DE  FREUDENREiCH.  De  la  resis- 
tance des  bacteries  aux  hautes  pressions 
combinees  avec  une  elevation  de  la  tem- 
perature. Annales  de  Microg.,  T.  iv,  1891, 
pp.  105-119. 

Milk  subjected  to  a  pressure  of  many  atmospheres 
(78-90)  for  several  hours  at  45°  to  63°  C  was  not  steril- 
ized. High  pressure  for  a  week  also  failed  to  sterilize  it. 

('93).  D'ARSONVAL  ET  CHARRIN.  Pression  et  microbes. 
La  semaiine  medicale,  1893,  T.  xin,  p.  251. 
Rev.  in  Centralb.  f.  Bakt.,  Bd.  xiv,  1893,  p. 
64.  See  also  C.  R.  des  se.  et  mem.  de  la 
soc.  de  Biol.,  Paris,  20  mai,  1893,  pp.  532-533. 

B.  pyocyaneus,  in  fresh  bouillon  cultures,  was  subjected 
to  a  pressure  of  50  atmospheres  under  carbon  dioxide. 
All  were  dead  inside  of  24  hours.  Kven  two  hours' exposure 
interfered  with  the  reproductive  function,  i.  e.,  lessened 
the  number  of  organisms  capable  of  producing  colonies, 
and  in  cultures  made  after  four  hours'  pressure  only 
traces  of  ability  to  form  pigment  remained.  In  cultures 
made  after  six  hours  exposure  there  was  no  formation 
of  pigmeut,  and  generally  no  colonies  when  sown  upon 
agar,  but  in  one  case  there  were  a  few. 

('94).  D'ARSONVAL  AND  CHARRIN.    See  xxxni. 
('94).  ROGER.     Action   des  hautes  pressions   sur  les 

microbes.     C.  R.  des  se.  de  '1'Acad.  des  sci., 

T.  cxix,  Paris,  p.  963. 

Pressures  ot  1,000  to  3,000  atmospheres  were  tried  with, 
out  destroying  the  bacteria.  Certain  functions,  however- 
were  destroyed,  e.  g.,  pathogenicity. 

('97).  MALFITANO,  G.  Sul  comportamento  dei  micro- 
organism! a'll'azione  dei  gasi  compressi. 
Boll,  della  Soc.  medioo-chirurgica  di  Pavia, 

1897.  Rev.  in  Centralb.  f.  Bakt.,  xxm  Bd., 

1898,  pp.  233-236. 


246 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


XXXIII.    Action  of  Heat  and  Cold  on  Bacteria. 

("75).  EIDAM,  EDUARD.  Die  Einwirkung  verschied- 
ener  temperaturen  und  des  Eintrocknens  auf 
die  Entwicklung  von  Bacterium  termo  Duj. 
Cohn's  Beitrage  z.  Biol.  d.  Pflanzen,  Bd.  I, 
Heft  3,  pp.  208-224,  Breslau,  1875. 

('77).  FRISCH,  A.  Ueber  den  Einfluss  niederer  Tem- 
peraturen auf  die  Lebensfahigkeit  der  Bac- 
terien.  Sitzungsber.  der  K.  Acad.  der 
Wissensch.  Wien.  Math.-natur.-wissenschaf- 
ten  Classe,  Mai,  1877,  Bd.  LXXV,  in  A'bt., 
pp.  257-269. 

('77).  TYNDALL,  JOHN.  On  heat  as  a  germicide  when 
discontinuously  applied.  Proc.  Roy.  Soc., 
London,  vol.  xxv,  1877,  No.  178,  pp.  569-570. 

('79)-  CHAMBERLAND,  CH.  Resistance  des  germes 
des  certains  o.rganismes  a  la  temperature  de 
100  degres;  conditions  de  leur  developpe- 
ment.  C.  R.  des  se.  de  1'Acad.  des  sci.,  T. 
i,xxxviil,  1879,  pp.  659-661. 

('82).  LEBEDEFF,  A.  Contribution  a  1'etude  de  1'action 
d«  la  chaleur  et  de  la  dessication  sur  la 
virulence  des  liquides  septique  et  sur  les 
Torganismes  inferieurs.  Archives  de 
Physiol.  normale  et  Path.,  Ser.  n,  T.  x,  pp. 
175-204,  1882. 

('84).  PICTET,  R.,  ET  YUNG,  E.  De  1'action  du  froid 
.sur  les  microbes.  C.  R.  des  se.  de  1'Acad. 
des  sci.,  T.  xcvm,  1884,  pp.  747-749. 

('87).  PRUDDEN.    See  XLVI. 

('87)  .  ESM  ARCH,  E.  Der  Henneberg'sche  Desinfector. 
Zeitschr.  f.  Hyg.,  Bd.  n,  1887,  pp.  342-368. 

('88).  GLOBIG.  Ueber  einen  Kartoffel-Bacillus  <mit 
ungewdhnlich  iwiderstandsfahigen  Sporen. 
Zeitschr.  f.  Hyg.,  Bd.  m,  1888,  pp.  322-332. 

('88).  GRUBER,  MAX.  Notiz  iiber  die  Widerstand- 
fahigkeit  der  Sporen  von  Bacillus  subtilis 
gegen  Wasserdampf  von  100°  C.  Centralb. 
f.  Bakt.,  1888,  Bd.  in,  pp.  576-577. 

In  six  tubes  of  hay  infusion  inoculated  with  spores  of 
B.  subtilis,  sealed  by  heating  neck  in  flame,  and  then 
steamed  %  hour,  there  was  an  abundant  growth  of  the 
hay  bacillus  in  36  hours  at  37°  C.  Subsequently  21  sam- 
ples of  spores,  dried  on  silk  threads  and  exposed  to 
streaming  steam  tor  2%  hours,  in  Thursfield's  apparatus, 
grew  readily  ;  in  24  hours,  at  35°  C.  ,  there  was  a  most  lux- 
uriant vegetation. 

('88).  FISCHER,    B.      Bakterienwachstum   bei    o°    C. 

Centralb.  f.  Bakt.,  Bd.  iv,  1888,  pp.  89-92. 
('90).  LUSTIG,  ALEXANDER.     Bin  rother  Bacillus  im 
Flusswasser.     Centralb.    f.    Bakt.,    Bd.   vni, 
1890,  pp.  33-40. 

Lustig  isolated  a  motile  bacillus  from  river  water, 
which  grew  from  room-temperature  (probably  15°  C  )  to 
60°  C. 

('92).  FORSTER,  J.    Ueber  die  Entwickelung  von  Bak- 
terien  bei  niederen  Temperaturen.    Centralb. 
f.  Bakt,  XH  Bd.,  1892,  pp.  431-436. 
The  kinds  of  bacteria  able  to  grow  at  o°  are  not  very 
numerous,  but  seem  to  be  widely  distributed,  especially 
in  water  and  on  the  surface  and  in  the  intestinal  tract  of 
fresh-water  fish  and  salt-water  fish. 

,  RAOUL.  De  1'ernplpi  methodique  des 
basses  temperatures  en  biologic.  Archiv.  d. 
sci.  phys.  et  nat.,  3e  Periode,  T.  xxx,  pp. 
293-314,  Geneve,  1893. 

Experiments  with  hlgheranimals  and  plants,  infusoria. 
microbes,  diatoms.  With  these  two  latter,  excessive  and 
prolonged  cold  gave  negative  results.  They  were  sub- 
jected to  a  temperature  of  minus  200°. 


('93).  PICTET,  RAOUL. 


('93).  PHYSALIX.  Influence  de  la  chaleur  sur  la  pro- 
priete  sporogene  du  Bacillus  anthracis. 
Abolition  persistante  de  cette  fonction  par 
heredite  des  characteres  acquis.  Arch,  de 
physiol.  nonmale  et  path.,  Paris,  1893,  T.  v, 
ser.  5,  pp.  217-225. 

('94).  D'ARSONVAL  ET  CHARRIN.  Influence  des  agents 
cosmiques  (electricite,  pression,  lumiere, 
froid,  ozone,  etc.)  sur  1'evolution  de  la  cell- 
ule bacterienne.  Ardi.  de  physiol.  normale 
et  path.,  1894,  T.  vi,  series  5,  pp.  335-342. 

('94).  WALDO  AND  WALSH.    See  XLVIII. 

('94).  HAVEMANN.  Ueber  das  Wachsthum  von 
Mikroorganismen  bei  Eisschranktemperatur. 
(Inaug.  Diss.)  8vo.,  21  pp.,  Rostock,  1894. 

Not  seen. 

('95).  STERNBERG,  GEORGE  M.  What  shall  be  the 
methods  followed  in  determining  the  rela- 
tion of  bacteria  to  temperature?  Jour.  Am. 
Public  Health  Asso.  Ann.  vol.  xx,  1895, 
pp.  411-414. 

(*95).  KLEPZOFF,  CONST.  Zur  Frage  iiber  den  Ein- 
fluss niederer  Temperaturen  auf  die  vegeta- 
tiven  Formen  des  Bacillus  anthracis.  Cen- 
tralb. f.  Bakt.,  xvii  Bd.,  1895,  pp.  289-295. 

Exposure  to  intense  cold  (average  —  24°  C.)  for  12  days 
killed  the  anthrax  organism  in  blood  and  various  organs. 
Exposure  for  25  days  (at  —  i°  to  —  24°  C. ,  average  —  10.40° 
C.)  killed  agar  cultures.  The  colonies  in  agar  plates  be- 
came less  and  less  numerous  as  time  passed  No  spores 
were  present.  L,ong  exposure  reduced  the  virulence. 

('95).  MIQUEL,  P.,  ET  LATTRAYE.  De  la  resistance 
des  spores  des  bacteries  aux  temperatures 
humides  egales  et  superieures  a  100°.  Ann. 
de  micr.,  T.  vn,  1895,  pp.  110-122,  158-170, 
and  205-218. 

('99).  DANNAPPEL,  MAX.  In  wie  weit  ist  die  hoherc 
Widerstandsfahigkeit  der  Baktoriensporen 
ein  allgemeines  Charakteristiikum  derselben 
gegenitber  den  vegetativen  Spaltpilzformcn? 
8vo.,  pp.  27.  Konigsberg,  i.  Pr.,  1899,  von 
E.  Karg  u.  R.  Manneck. 

Some  spores  are  said  to  show  only  a  slight  resistance  to 
steam  at  99°  C.  Of  25  species  obtained  from  soils,  decay- 
ing  mixtures,  milk,  butter,  etc.,  and  said  to  be  sporifer- 
ous,  all  but  three  were  destroyed  by  exposure  to  steam  at 
99°  C.  for  10  minutes  and  all  but  8  by  exposure  for  5  minutes 
while  4  were  killed  by  exposure  for  as  short  a  time  as  15, 
seconds,  and  two  others  by  exposure  for  i  minute.  The 
names  of  these  organisms  are  not  given,  so  that  the  ex- 
periments cannot  be  duplicated,  and  in  most  cases  it  is 
not  stated  that  the  sporiferous  nature  of  these  bacteria 
was  settled  definitely  by  seeing  the  spores  germinate.  It 
is  possible,  therefore,  that  some  of  the  extremely  sensi- 
tive forms  were  not  actually  spore-bearing,  but  only  gave 
microscopic  appearances,  which  were  interpreted  as  such. 
It  is  possible,  also,  that  the  spores  were  tested  before  they 
were  fully  matured.  Onlytwoofthe  very  sensitive  forms 
were  examined  critically,  owing,  it  is  said,  to  lack  of 
time,  and  of  one  of  these  sensitive  forms  it  is  snid  :  "A 
direct  observation  of  the  germination  was  not  undertaken 
because  the  spore  nature  of  the  culture  appeared  unques- 
tionable." Even  heating  for  i  to  3  minutes  at  75°  to  80°  C 
destroyed  this  organism.  Germination  of  the  other  was 
observed.  The  maximum  temperature  which  could  be 
endured  in  this  case,  for  I  minute,  was  75°  C-  Both  were 
green  spores.  Both  were  double  stained  by  Moeller's 
method. 

('99).  RAVENEL,  M.  P.  The  resistance  of  bacteria  to 
cold.  New  York  Medical  News,  vol.  LXUV, 
1899.  Also  a  separate,  5  pp.  Rev.  in  Cen- 
tralb. f.  Bakt.,  xxvni  Bd.,  1900,  p.  751. 

Tests  in  liquid  air :  B.  diphtheria  was  alive  at  end  of  30 
minutes,  B.  typhi  and  B .  prodigiosus  at  the  end  of  60 
minutes,  B.  anthracis  after  3  hours. 


ACTION    OF    HEAT    AND    COLL);    THERMOPHILIC    BACTERIA. 


247 


('99).  SMITH.  THEOBAIA  The  thermal  death  point 
of  tubercle  bacilli  in  milk  and  some  other 
fluids.  Journal  of  Experimental  Med.,  vol. 
iv,  1899.  pp.  217-233.  Rev.  in  Central!),  f. 
Bakt.,  .\.\vin  Bd.,  1900,  p.  409. 

When  embedded  in  the  film  on  the  surface  of  milk,  Dr. 
Smith  found  the  tubercle  organism  resisted  a  tempera- 
ture of  60°  C.  for  an  hour. 

(V.y).  KASANSKY,  M.  W.  Die  Einwirkung  der  Win- 
terkalte  auf  die  Pest-  und  Dip'htheriebacillen. 
Central!),  f.  Bakt.,  xxv  Bd.,  1899,  pp.  122-124. 

These  organisms  withstood  exposure  for  6  mouths  to 
severe  temperatures.  They  weie  frozen  all  of  the  time 
for  the  first  five  months.  From  December  4  to  28,  aud 
again  from  February  13  to  March  9,  the  maximum  tem- 
perature was  — 10°  to— 234°  C,  and  the  minimum  was 
—  14°  to  — 33.8°  C. 

('•.,9).  LEVIN.     See  xui. 

('99).  MIKONESCO,  THEODOR  G.  Ueber  eine  besondere 
Art  der  Beeinfltissung  von  Mikroorganis- 
men  durch  die  Temperatur.  Hygien.  Rund- 
schau, Jahrg.  ix,  1899,  pp.  961-964.  Rev.  in 
Centralb.  f.  Bakt.,  xxvn  Bd.,  1900,  p.  86. 

('oo).  MEYER,  J.  Ueber  Einwirkung  fliissiger  Luft 
auf  Bakterien.  Centralb.  f.  Bakt.,  xxvm 
Bd.,  1900,  pp.  594-595- 

Anthrax  spores  and  Staphylococcus  pyog.  aureus  were 
tested.  The  exposure  to  the  liquid  air  varied  from  5 
seconds  to  15  minutes.  Neither  organism  was  killed. 
The  temperature  of  liquid  air  is  190°  to  — 220°  C  .accord- 
ing to  Spiess,  and  — 182°  to  —  192°  C.  according  to  Mac- 
faclyen 

C'oo).  SEDGWICK,  W.  T.,  AND  WINSLOW,  C.  E.  A. 
Experimental  and  statistical  studies  on  the 
influence  of  cold  upon  the  bacillus  of  typhoid 
fever,  and  its  distribution.  Jour.  Bost.  Soc. 
Med.  Sci.,  vol.  iv,  No.  7,  1900,  pp.  181-182. 
See  also  Centralb.  f.  Bakt.,  xxvn  Bd.,  1900, 
p.  684. 

30  to  60  per  cent  of  the  bacilli  were  destroyed  in  water 
during  the  first  hour  ot  freezing.  After  exposure  for  two 
weeks  99  per  cent  were  destroyed.  "  The  last  two  or  three 
germs  per  thousand  appear  to  be  very  resistant,  some 
remaining  a'ter  twelve  weeks  of  freezing.  The  four  races 
used  showed  constant  individual  differences  in  their  sus- 
ceptibility to  cold.  Alternate  freezing  aid  thawing  was 
tested  and  found  only  slightly  more  destructive  than  con- 
tinuous freezing."  As  several  races  of  typhoid  organism 
were  tested,  we  may  infer  that  ice  is  not  very  likely  to 
communicate  typhoid  fever. 

('oo).  PARK,  WM.  HALLOCK.  A  few  experiments 
upon  the  effects  of  low  temperature  and 
freezing  on  typhoid  bacilli.  Jour.  Bost.  Soc. 
Med.  Sci.,  vol.  iv,  No.  8,  1900,  pp.  213-216. 

Cultures  were  used  from  twenty  different  cases  of 
typhoid  fever.  They  behaved  when  frozen  much  as  Sedg- 
wick  and  Winslow's.  On  the  average,  at  the  end  of 
twelve  weeks'  freezing  only  0.05  of  one  per  cent  remained 
alive,  i  e. ,  1,250  per  cubic  centimeter  as  against  2,560,410 
per  cubic  centimeter  at  the  beginning. 

"At  twelve  weeks  the  bacilli  in  the  ice  from  nine 
sources  are  all  dead.  Two  more  show  no  growth  in  i  cc. 
The  others  contain  from  80  to  11,000  in  each  cc.  of  ice. 
Only  one,  however,  contains  over  1,000  ^culture  9). 
When  typhoid  bacilli  are  in  feces,  freezing  does  not 
exert  so  much  of  an  effect.  Thus  typhoid  and  colon 
bacilli,  originally  37,000  to  a  loopful  of  feces,  wrre  still 
12,000  at  the  end  of  five  weeks'  exposure  to  a  temperature 
ranging  daily  between  zero  and  28°  F. ,  and  typhoid 
bacilli  as  well  as  colon  were  still  abundant  in  the  feces  at 
nine  weeks.  It  is  a  difficult  malterto  say  for  just  how 
long  a  period  ice  made  from  infected  water  remains  dan- 
gerous. The  bacilli,  even  when  few  in  number,  are  often 
vigorous  and  fully  virulent,  and,  so  far  as  I  am  aware,  we 
are  ignorant  as  to  the  number  of  bacilli  required  to  start 
infection  in  man.  The  longer  the  infected  ice  remains 
frozen  the  less  the  number  of  pathogenic  bacteria  which 
remain  alive  in  it." 


Coi).  PARK,  W.  H.  Duration  of  life  of  typhoid 
bacilli,  derived  from  twenty  different 
sources,  in  ice.  Abstract  of  paper  read  at 
2d  meeting  Soc.  Am.  Bacteriologists,  Dec., 

1900,  Centralb.  f.  Bakt.,  i  Abt.,  Bd.  xxix, 

1901,  pp.  444-445- 

This  describes  the  completion  of  an  experiment  already 
reported  upon  in  part  (see  above).  At  the  end  of  the 
twenty-second  week  of  exposure  the  bacilli  were  dead  in 
all  the  cultures  of  each  one  of  the  twenty  races  tested  by 
freezing. 

Coi).  D'ARSONVAL.      La   pression   osmotique    et    son 
role    de    defense    centre    le    froid    dans    la 
cellule  vivante.    C.  R.  des  se.  de  1'Acad.  des 
sci.,  Paris,  1901,  T.  cxxxm,  pp.  84-86. 
The  fluid  in  the  bacteria  is  probably  not  solidified,  if 
the  cell  is  not  ruptured,  owing  to  the  enormous  osmotic 
pressure  in  those  small  organisms.     By  lowering  the  os- 
motic tension  the  author  thinks  that  any  cell  may  be 
killed  by  cold. 

('02).  SCHMIDT-NIELSEN.  SIGVAL.  Ueber  einige  psy- 
chrophile  Mikroorganismen  und  ihr  Vor- 
kommen.  Centralb.  Bakt.,  Abt.  2,  Bd.  ix, 

1902,  pp.  145-147. 

(oa).  MACFADYEN,  ALLEN,  AND  ROWLAND,  SYDNEY. 
On  the  suspension  of  life  at  low  tempera- 
tures. Abstract  of  paper  read  before  Sec- 
tion K  of  the  British  Association,  Belfast, 
1902.  Annals  of  Botany,  vol.  xvi,  1902  pp 
589-590. 

Various  bacterial  organisms  were  exposed  from  20  hours 
to  7  days  at —190°  C.  "These  exposures  did  not  produce 
any  appreciable  impairment  in  the  vitality  of  the  organ- 
isms, etc."  Also  10  hours  at  —252°  C.  the  temperature  of 
liquid  hydrogen  had  no  appreciable  effect  on  the  vitality 
of  the  micro-organisms  tested.  Bacillus  typhosus,  B.coll- 
communis,  Staphylococcus  pyogenes  aureus  and  a  Sach- 
aromycete  grew  after  exposure  to  liquid  air  for  six 
mouths.  "In  no  instance  could  any  impairment  of  the 
vitality  of  the  organisms  be  detected." 

The  objection  to  these  statements  is  that  quantitative 
determinations  appear  not  to  have  been  made,  at  least 
there  is  no  mention  ot  any.  The  writer  of  this  review 
obtained  a  decided  diminution  of  the  number  of  viable 
bacteria  in  several  species  by  exposure  to  liquid  air  for  20 
hours. 

('02).  MACFADYEN,  ALLEN.  On  the  influence  of  the 
prolonged  action  of  the  temperature  of 
liquid  air  on  micro-organisms,  and  on  the 
effect  of  mechanical  trituration  at  the  tem- 
perature of  liquid  air  on  photogenic  bacteria. 
London,  Proc.  R.  Soc.,  vol.  LXXI,  No.  468, 
Oct.,  1902,  pp.  76-77. 

"  The  above  experiments  show  that  a  prolonged  expo- 
sure of  six  mouths  to  a  temperature  of  about  —  190°  C.  has 
no  appreciableeffect  on  the  vitality  of  micro-organisms." 

The  organisms  tested  were  H.  lyi'ho*ui,Jl.  coll  corn- 
munis.  Staphylococcu*  pynaenes  attreus,  and  a  yeast. 
The  triturated  bacteria  lost  their  luminosity. 

('05).  SMITH  AND  SWINGLE.    See  p.  83. 


XXXIV.    Thermophilic  Bacteria. 

(>9).  MIQUEL,    P.      Title?      Bull,    de    la    statistique 

municipale  de  Paris,  Decembre,  1879. 
Not  seen. 

He  discovered  in  the  water  of  the  Seine  an  immobile, 
rod-shaped  Schizomycete  capableof  living  and  develop- 
ing at  the  temperature  of  70°  C. 

('81).  VAN    TIEGHEM,    PH.      Stir    des    bacteriacees 
vivant  a  la  temperature  de  74°  C.     Bull    Soc 
bot.  de  France,  T.  28,  1881,  pp.  35-36. 
This  author  cultivated  several  species  of  thennophilic 
bacteria  at  70°  C.,  and  some  at  higher  temperatures. 

C8r).  MIQUEI..     Thermobacteria.     Annuaire  de  1'Ob- 
servatoire  de  Montsouris,  pour  1881,  p.  464. 


248 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('86).  CERTES,  A.,  ET  GARRIGOU.  De  la  presence  con- 
stante  de  micro-organismes  dans  ^les  eaux 
de  Luchon,  recueillies  au  griffon  a  la  tem- 
perature de  64°,  et  de  leur  action  sur  la 
production  de  la  baregine.  C.  R.  des  se.  de 
1'Acad.  des  sci.,  T.  cm,  I&S6,  pp.  703-706. 

('88)  GLOBIG  Ueber  Bakterien-Wachsthum  bei  50 
bis  70°.  Zeitschr.  f.  Hyg.,  Bd.  in,  1888,  pp. 
294-321.  Rev.  in  Centralb.  f.  Bakt.,  Bd.  in, 
1888,  pp.  366-368. 

Globig  obtained  30  sorts  of  bacteria  which  grew  on 
potato  at  58°  C.  At  68°  only  a  few  of  them  continued  to 
grow.  At  70°  C.  there  were  only  scattering  colonies,  and 
at  higher  temperatures  there  was  no  growth.  These 
organisms  were  not  pathogenic  to  mice.  As  a  rule, 
growth  began  at  about  50°  C.,  i.  e.,  about  13  degrees  above 
blood-heat.  One  would  not  grow  at  37°  C.  or  50°  C.,  but 
grew  at  60°.  One  grew  all  the  way  from  15°  or  20"  C.  to 
68°  C  Spores  were  often  formed  in  24  hours.  None  of 
these  were  from  feces  or  sewage.  Most  were  bacilli 
(rods). 
('88).  MIQUEL,  P.  Monographic  d'un  baoille  vivant 

a    au-dela    de    70°    centigrades.     Ann.    de 

micro.,  1888,  T.  I,  pp.  3-10. 

This  organism  will  not  grow  at  temperatures  under  40°, 
nor  above  72°  C.  Its  optimum  temperature  is  65°  to  70°  C. 

('90).  COHN,  FERDINAND.  Ueber  Warmeerzeugung 
durch  Schimmelpilze  und  Bakterien.  Vor- 
trag.,  Breslau,  1890. 

('93).  COHN,  F.    Ueber  therniogene  Bakterien.    Ber. 
d.    Deutsch.   bot.    Gesellsch.,    Bd.    xi,    1893, 
Gen.  Versarnlungs-Heft,  pp.  66-69. 
Conn  found  that  when  cotton  wool  waste  was  moist- 
ened it  reached  a  temperature  of  67.2°  C.  in  24  to  36  hours, 
and  then  slowly  cooled  (6  days)  to  the  air-temperature. 
When  the  same  waste  was  sterilized  there  was  no  rise  in 
temperature. 

('94).  MACFADYEN,  ALLAN,  AND  BLAXALL,  FRANK  R. 
Thermophilic  bacteria.  Journal  of  Path- 
ology and  Bacteriology,  vol.  in,  1894,  pp. 
87-99.  See  also  Br.  Med.  Jour.,  No.  1,760, 

1894,  p.  644. 

These  authors  obtained  from  garden  soil  an  abundant 
growth  of  thermophilic  bacteria  on  agar  at  60°  to  65°  C, 
They  also  isolated  these  organisms  from  feces,  sewage, 
sea-water,  dirt  of  London  streets,  Thames  water,  Thames 
mud,  straw,  surface  soil,  and  soil  5  feet  4  inches  down. 
These  thermophilic  bacteria  are,  therefore,  most  widely 
distributed.  There  were  guile  a  variety  of  species— at  least 
twenty.  All  were  bacilli ;  all  were  spore-bearing.  Some 
were  actively  motile.  Active  motility  continued  for  three 
weeks  in  one  hanging  drop.  The  colonies  developed  very 
rapidly  in  agar-plates.  Fifteen  sorts  were  tested  as  fol- 
lows :  None  of  them  grew  at  22^  or  at  37°  C.  Horse-dung 
organisms  grew  at  40°  to  42°  C.  Six  sorts  grew  slowly  at  50° 
to  52°  C.,  and  more  abundantly  at  60°  to  6s0  C.  Two  grew 
first  at  56°,  and  four  refused  to  grow  at  56°,  but  grew  when 
the  temperature  was  raised  to  60°  C.  None  would  grow 
at  75°.  The  lower  limit  of  growth  for  nearly  all  was  50° 
C.  and  the  upper  near  75°  C.  Boiling  for  ten  minutes  did 
not  destroy  these  organisms.  The  optimum  temperature 
for  growth  is  said  to  be  60°  to  65°  C.  These  organisms  did 
not  ferment  sugars  and  did  not  thrive  on  substrata  con- 
taining sugars,  these  substances  seeming  to  retard  growth. 
One  changed  starch  to  sugar. 

Query  :  How  do  these  organisms  exist  in  a  climate  as 
cold  as  that  of  Edinburg  ? 

('95).  RABINOWITSCH,  LYDIA.  Ueber  die  thermo- 
philen  Bakterien.  Zeitschr.  f.  Hyg.,  Bd.  xx, 

1895,  pp.  154-164.    Leipsic. 

These  thermophilic  organisms  were  found  in  snow,  in 
earth,  in  the  dust  of  a  street  in  Berlin.  They  were  very 
abundant  in  the  water  of  the  Spree  (7,000  to  8,000  per  cubic 
centimeter).  They  are  abundant  in  the  dung  of  horses  and 
cows,  and  also  more  or  less  so  in  the  excrement  of  goats, 
rabbits,  guinea  pigs,  dogs,  mice,  doves,  hens,  ducks,  par- 
rots. They  occur  in  the  whole  digestive  tract  ot  man.  and 
are  found  in  certain  fish,  frogs,  and  some  other  cold- 
blooded animals.  Miss  R.  also  found  them  abundant  in 
germinating  barley  in  a  brewery.  They  occur  also  in  milk, 
even  after  it  is  boiled.  She  isolated  and  studied  8  species. 
All  produced  spores.  None  were  pathogenic  to  mice  or 


doves.  The  highest  temperature  at  which  any  of  them 
would  grow  was  75°  C.,  and  growth  at  this  high  tempera- 
ture was  slight.  There  was  an  abundant  growth  at  58°  to 
68°  C.,  and  the  optimum  is  said  to  be  60°  to  70°.  They  are 
very  resistant  to  moist  heat  and  also  to  dry  heat.  They 
were  not  killed  by  exposure  to  streaming  sleam  for  5  to  6 
hours.  While  growing  best  at  high  temperatures,  these 
organisms  can  grow  slowly  facultative-anaerobically  at 
33°  to  40°  C.,  and  the  author  thinks  that  they  multiply  in- 
side warm-blooded  animals.  She  found  the  temperature 
of  dung-piles  as  high  as  62°  to  66°  C. 

('95).  KARLINSKY,  JUSTYN.  Zur  Kenntniss  der  Bac- 
terien  der  Thermalquellen.  Hygienische 
Rundschau,  1895,  Bd  v,  pp.  685-689. 

('96).  TEICH,  M.  Beitrag  zur  Kenntniss  thermo- 
philer  Bacterien.  Hygienische  Rundschau, 
1896,  Bd.  vi,  No.  22,  pp.  1,094-1,095. 

('98).  LAXA,  O.     Ueber  einen  thermophilen  Bacillus 
aus     Zucker-Fabriksproduoten.       Vorlaufige 
Mittheilung.    Zeitschr.  f.  Zuckerindustrie  in 
Bdhmen,  Bd.  xxn,  1898,  p.  376. 
Not  seen. 

('98).  OPRESCU.  Studien  iiber  thermophile  Bakterien. 
Arch.  f.  Hyg.,  Bd.  xxxm,  1898,  p.  164.  Rev. 
in  Centralb.  f.  Bakt.,  Bd.  xxv,  1899,  p.  360. 
('99).  TSIKLINSKY,  MLI.E.  Sur  les  microbes  ther- 
mophiles  des  sources  thermales.  Ann.  de 
1'Ins.t.  Pasteur,  T.  xni,  1899,  pp.  788-795- 
Bibliog.  of  13  titles. 

The  author  isolated  five  thermophilic  organisms  from 
hot  springs.  One  form  grew  readily  at  70°  C. 

('99).  MICHAELIS,  GEORG.  Beitrage  zur  Kenntniss 
der  therniophilen  Bakterien.  Arch.  f.  Hyg., 
Bd.  xxxvi,  Hft.  3,  1899,  pp.  285-293.  Rev. 
in  Centralb.  f.  Bakt.,  xxvn  Bd.,  1900,  p.  537. 

Describes  and  names  four  new  thermophilic  organisms. 
All  have  three  specific  names,  and  one  has  four,  to-wit  : 
Bacterium  thermophilus  aquatilis  liquefaciens  aerobius. 

('99).  CAMBIER.     Thermophilic  bacteria  as  ferments, 
action  on  glucose.    Rev.  de  phys.  et  de  chim., 
1899,  p.  223. 
Not  seen. 

('02).  RUSSELL,  H.  L-,  AND  HASTINGS,  E.  G.  A 
Micrococcus,  the  thermal  death  limit  of 
which  is  76°  C.  Centralb.  f.  Bakt.,  2  Abt., 
Bd.  vin,  1902,  pp.  339-342,  i  plate. 

('03).  SETCHELL,  WM.  A.  The  upper  temperature 
limits  of  life.  Science,  n.  s.,  vol.  xvn,  1903, 

PP-  934-937- 

Hot  springs  were  studied  in  three  localities  in  Cali- 
fornia and  in  Yellowstone  National  Park.  Author  found 
only  CyanophyceaJ  and  Bacteria  in  strictly  thermal  waters. 
The  Cyanophycese  were  found  at  65°  to  68°  C.,  and  spar- 
ingly up  to  75"°  to  77°  C.  The  bacteria  were  abundant  at 
70°  to  71°,  and  occurred  in  considerable  quantity  at  82°  C. 
and  89°  C.  "The  temperature  of  89°  C.  is  the  highest  at 
which  I  have  been  able  to  find  any  organisms  Hying." 
Care  was  taken  to  determine  the  temperatures  in  the 
exact  places  frequented  by  the  organisms,  so  as  to  remove 
the  objection  that  lies  against  many  of  the  earlier  obser- 
vations. 


XXXV.    Resistance  to  Dry  Air. 

('75).  EIDAM.    See  xxxm. 

('92).  MOMONT,  L.  Action  de  la  >dessication,  de  1'air, 
et  de  la  lumiere  sur  ila  baeteridie  char- 
ilxmneifse  nlamenteuse.  Ann.  de  1'Inst.  Pas- 
teur, 1892,  T.  vi,  pp.  21-31. 
SWAN,  ALLEN  P.  On  the  resisting  vitality  of 
the  spores  of  Bacillus  megaterium  to  the 
condition  of  dryness.  Annals  of  Botany, 
vol.  vn,  p.  153-154,  1893- 


('93). 


RESISTANCE  TO  DRY  AIR;  ACTION  OF  ACIDS  AND  ALKALIES,  ETC.         249 


('94).  WALLICZEK,  HEINRICH.  Die  Resistenz  des 
Bacterium  coli  commune  gegen  Eintrock- 
nung.  Centralb.  f.  Bakt.,  Bd.  XV,  1894,  pp. 
949-950. 

B.  coli  proved  quite  sensitive  to  dry  air.  The  writer  of 
this  abstract  has  found  great  differences  among  bacteria 
e.  g.  B.  tracheiphilus  was  killed  by  a  few  minutes'  expos- 
ure ou  cover-glasses,  while  Bact.  hyacinthi  lived  under 
similar  conditions  for  more  than  a  month.  Jones  found 
his  Bacillus  carotovorus  to  be  even  more  sensitive  than 
B.  tracheiphilus.  See  following  citations. 

C'9S)-  SMITH,  ERWIN  P.     Bacillus  tracheiphilus,  etc. 

Centralb.  f.  Bakt.,  2  Abt.,  Bd.  i,  p.  ,370. 
('97).  MIQUEL,  P.     Sur  la  longevite  des  germes  des 

bacteries  dans  les  poussieres  et  dans  le  sol. 

Ann.  de  micr.,  1897,  T.  ix,  pp.  199-207  and 

251-259. 
Coi).  JONES,  L.  R.     A  soft  rot  of  die  carrot,  etc. 

I3th  Ann.  Rep.  Vt.  Agric.  Exp.  Sta.  for  1900. 

Burlington,  Vt.,  1901.    Sec  p.  328  for  refer- 
ence to  this  subject. 
('01).  SMITH,  ERWIN  P.    The  cultural  characters  of 

Ps.  hyacinthi.  etc.     Bull.  28,  Div.  Veg.  Phys. 

and   Path.,   U.    S.   Dep.   Agr.,   Washington, 

D.  C.,  1901,  p.  145. 


XXXVI.     Action  of  Acids  and  Alkalies. 

('86).  ABBOTT,  A.  C.  The  germicidal  value  of  some 
of  the  vegetable  acids.  The  Medical  News, 
Phil  a.,  1886,  9  Jan.,  pp.  33-34. 

('92).  DELBRUECK,  M.  Ueber  das  Verhalten  der 
Cholerabacillen  auf  frischen  Friichten, 
einigen  Genuss-  und  Nahrungsmitteln.  Son- 
derabdruck  aus  den  Veroffentlichungen  des 
Kaiserlichen  Gesundheitsamte,  1892,  No.  42, 
vom  19  October,  Berlin.  Verlag  von  Julius 
Springer,  1892,  pp.  812-824. 

The  per  cent  of  malic  acid  in  many  fruits  is  given.  This 
varies  from  0.13  (certain  pears)  to  2.65  (red  currantsV  In 
the  feebly  acid  fruits,  the  cholera  bacilli  were  dead  inside 
of  3  to  7  days  :  in  the  the  tarter  fruits  thev  retained  their 
vitality  only  for  a  period  measured  by  hours.  Usually 
they  were  dead  in  from  i  to  6  hours. 

('92).  SCHLUETER,  G.  Das  Wachstum  der  Bakterien 
auf  saurem  Nahrboden.  Centralb.  f.  Bakt., 
Bd.  xi,  1892,  pp.  589-598. 

A  dozen  different  bacteria  were  tested  in  "  ordinary  nu- 
trient gelatin  "  and  in  fish  glue,  with  addition  of  lactic 
acid,  aium,  tartaric  acid,  citric  acid,  acetic  acid,  and  hy- 
drochloric acid.  Several  organisms  tolerated  as  much 
as  i  per  cent  of  lactic  acid,  or  i  per  cent  tartaric  acid, 
but  their  growth  was  slow  and  usually  feeble.  Several 
grew  feebly  in  the  presence  of  l/t  per  cent  alum.  Six 
grew  abundantly  in  gelatin  acidified  with  citric  acid. so 
that  H  cc.  of  the  gelatin  required  for  its  neutralization 
4  cc.  of  sodium  carbonate  water  of  the  strength  5.3:1000. 
In  fish  glue  containing  0.15  per  cent  acetic  acid,  several 
grew,  but  only  feebly.  Six  grew  in  fish  glue  containing 
0.075  per  cent  hydrochloric  acid.  The  anthrax  organism 
grew  better  with  0.2  per  cent  alum  than  on  a  neutral 
substratum. 

(93).  HESSE,  W.  Ueber  den  Einfluss  der  Alka- 
lescenz  des  Nahrbodens  auf  das  Wachsthum 
der  Bakterien.  Zeitschr.  f.  Hyg.,  Bd.  xv, 
1893,  PP.  183-191- 

('9,3).  VOCES,  O.  Ueber  das  Wachstum  der  •Cholera- 
bacillen auf  Kartoffeln.  Centralb.  f.  Bakt., 
Bd.  xm,  1893,  pp.  543-550. 

Organism  woulci  uot  grow  on  potato  as  ordinarily  pre- 
pared, but  grew  well  at  37°  C.  (and  more  slowly  at  20°)  on 
the  addition  of  a  2to  3  percent  solution  of  sodium  chlor- 
ide. Nearly  as  good  results  were  obtained  with  %  to  % 
per  cent  sodium  carbonate  solution.  Growth  was  also 
obtained  on  potato  with  J4  to  %  per  cent  sodium  hydrate 
solution. 


('97).  DEELEMAN,  M.  Der  Einfluss  der  Reaktion  des 
Nahrbodens  auf  das  Bakterienwachstum. 
Arbeit,  aus  dem  Kaiserl.  Gesundheitsamte, 
Bd.  xin,  1897,  Heft  3.  Rev.  in  Centralb.  f 
Bakt,  xxn  Bd.,  1897,  pp.  355-356. 

(98).  FERMI,  CLAUDIO.  Die  Mineral-  und  organ- 
ischen  Sauren,  die  Alkali,  die  Alkaloide,  das 
Jodkali  und  das  arsensaure  Kali  zur  Differ- 
enziierung  der  Mikroorganismen.  Centralb. 
f.  Bakt.,  Bd.  xxin,  1898,  pp.  208-217  and 
266-273. 

Of  the  plant  acids,  oxalic  was  found  to  be  the  most 
deleterious  to  the  Schizomycetes.  The  conclusions  are 
given  on  p.  266  et  seq. 


('96) 
('97) 

('97) 

('97) 

('98) 
('99) 
('99) 
('99) 

('99) 
Coo) 

Coo) 
Coo) 


XXXVII.    Agglutination  and  Precipitation. 

.  WIDAI,,  FERNAND.  Sero^diagnostic  de  la  fievre 
typhoide.  Bull,  et  mem.  de  la  soc.  med.  des 
hop.  de  Paris,  26  juin,  1896,  pp.  561-566. 

.  WIDAL,  P.,  ET  SICARD,  A.  Etudes  sur  le  sero- 
diagnostic  et  sur  la  reaction  agglutinante 
chez  les  typhiques.  Ann.  de  1'Inst.  Pasteur, 
T.  xi,  1897,  pp.  353-432. 

.  FLEXNER,  S.  A  recently  discovered  property 
of  the  blood  .serum  in  animals  immune  from 
certain  diseases,  and  its  application  to  the 
diagnosis  of  those  diseases  in  human  beings. 
Science  (n.  s.),  vol.  v,  pp.  193-194,  1897. 

.  MALVOZ,  E.  Reoherches  sur  '1'agglutination  du 
Bacillus  typhosus  .par  des  substances 
chimique.  Ann.  de  1'Inst.  Pasteur,  T.  xi, 
J897,  pp.  582-590. 

.  NICOLLE,  CHARLES.  Recherches  sur  la  sub- 
stance agglutinee.  Ann.  de  1'Inst.  Pasteur,, 
T.  xii,  1898,  pp.  161-191. 

.  BORDET,  JULES.  Le  mechanisme  de  1'agglutina- 
•tion.  Ann.  de  1'Inst.  Pasteur,  T.  xm,  1899, 
pp.  225-250. 

.  KRAUS,  R.  Ein  Beitrag  zur  Kenntniss  des 
Mechanismus  der  agglutination.  Wiener 
Klin.  Wochenschr.  1899,  Jahrg.  xn,  pp.  1-4. 

.  GRUBER.  Zur  Theorie  der  Agglutination. 
Munch,  med.  Wochenschr.,  1899,  No.  41. 
Rev.  in  Centralb.  f.  Bakt.,  xxvn  Bd.,  igoo, 
pp.  285-286. 

.  SABRAZES  ET  BRENGUES.  Agglutinines  chim- 
iques.  C.  R.  de  la  Soc.  de  biol.,  1899,  No. 
35.  P-  930.  Rev.  in  Centralb.  f.  Bakt.,  xxvn 
Bd.,  1900,  p.  756. 

.  ZIKES.  Ueber  das  Ausschleudern  von  Mikro- 
organismen unter  Zuhilfenahme  von  Fal- 
lungsmitteln.  Oesterr.  Chemiker-Zeitung, 
1900,  No.  2.  Rev.  in  Centralb.  f.  Bakt., 
xxvii  Bd.,  1900,  p.  628. 

.  SMITH,  R.  GREIG.  The  flocculation  of  bacteria. 
The  mechanism  of  agglutination.  Proceed- 
ings of  Linn.  Soc.  of  New  South  Wales, 
1900,  Part  i,  pp.  65-74,  75-83.  Also  a  sepa- 
rate (issued  Aug.  8,  1900). 

.  DURHAM,  HERBERT  E.  Some  theoretical  con- 
siderations upon  the  nature  of  agglutinins, 
together  with  further  observations  upon 
Bacillus  typhi  abdominalis,  Bacillus  enteri- 
tidis,  Bacillus  coli  communis,  Bacillus  lactis 
aerogenes,  and  some  other  bacilli  of  allied 
character.  Jour,  of  Exp.  Med.,  vol.  v,  pp. 
353-388. 


250 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('01).  WILSON,  ROBERT  J.  Observations  contributing 
to  precision  in  the  use  of  the  Widal  test  for 
typhoid.  N.  Y.  Univ.  Bull,  of  the  Med.  Sci., 
vol.  i,  No.  2,  1901,  pp.  87-92. 

('or).  DODGE,  CHARLES  WRIGHT.  A  short  method 
for  the  Widal  test.  Jour,  of  Applied  Micro., 
vol.  iv,  1901,  p.  1,565.  Also  a  separate. 

('02).  NEUFELD,  F.  Ueber  die  Agglutination  der 
Pncumokokken  und  iiber  die  Theorieen  der 
Agglutination.  Zeitschr.  f.  Hyg.,  1902,  Bd. 
XL,  pp.  54-72. 

('02).  EISENBERG,    PHILIPP,    UND    VOLK,    RICHARD. 
Untersuchungen  iiber  Agglutination.   Zeitsch. 
f.  Hyg.,  1902,  Bd.  XL,  pp.  155-195- 
Bibliog.  of  56  titles. 

('02).  Joos,  A.  Untersuchungen  iiber  den  Mechan- 
ismus  der  Agglutination.  Zeitsch.  f.  Hyg., 

1902,  Bd.  XL,  pp.  203-230. 

('02).  SMITH,  R.  GREW,.  Further  remarks  upon  the 
mechanism  oi  agglutination.  Proc.  Linnean 
Soc.  of  New  South  Wales,  vol.  xxvn,  1902, 
Part  i,  pp.  66-72.  Also  a  separate  (issued 
Aug.  22,  1902). 

('03).  FLEXNER,  SIMON.  An  aspect  of  modern  path- 
ology. Science,  n.  s.,  vol.  xvin,  No.  444, 

1903,  PP-  3-15- 


XXXVIII.    Antiseptics  and  Germicides. 

(See  also  XXXVI.) 

('70).  LISTER,  JOSEPH.  On  the  effects  of  the  anti- 
septic system  of  treatment  upon  the  salu- 
brity of  a  surgical  hospital.  Edinburgh, 
Edmonston  and  Douglas,  1870,  pp.  19. 

('74).  DAVAINE,  C.  Reoherches  relatives  a  1'action 
des  substances  antiseptiques  sur  le  virus  de 
la  septicemie.  Gaz.  med.  de  Paris,  1874,  p. 
44.  Reprinted  in  1'Oeuvre  de  Davaine, 
Paris,  1889. 

('75)-  LEWIN,  L.  Das  Thymol  ein  Antisepticum  mid 
Antifermentativum.  Virchow's  Archiv., 
Bd.  LXV,  1875,  pp.  164-189.  Polli's  Annali 
di  chimica  applicata  alia  med.  Milano,  vol. 
LXII,  1876,  pp.  321-324. 

('77).  POLLI,  G.  Sulle  proprieta  anti fermentative 
dell'acido  boracico  e  sue  applicazioni  alia 
terapia.  Mem.  1st.  Lomb.,  vol.  xm,  pp.  453- 
468.  Journ.  de  Pharm.,  et  de  chimie,  T. 
xxvi,  1877,  4  se.,  pp.  77-79. 

('79).  BOVET,  V.  Ueber  die  antiseptischen  Eigen- 
sohaften  der  Pyrogallussaure.  Journ.  f. 
prakt.  Chem.  Neue  Folge,  Bd.  xix,  pp. 
445-461,  1879. 

('80).  PAVESI,  C.  Del  solfato  di  potassa,  e  special- 
mente  della  sua  proprieta  antisettica,  anti- 
fermentativa.  Polli.  Annali,  vol  LXXI,  serie 
3a,  1880,  pp.  110-115. 

C8o).  ENDEMANN,  H.  Boracic  acid  as  a  preservative. 
Chem.  News,  vol.  XLI,  pp.  152-153,  1880. 

('80).  SCHWARTZ,    NICOLAI.      Ueber    das    Verbal-ten 
eini-ger    Antisettica    zu    Tabacksinfusbacte- 
rien.    Pharm.  Zeitschr.  f.  Russhnd,  Bd.  xix, 
1880,  pp.  610-625,  641-658,  673-685. 
Tested  chloroform  and  found  it  of  little  worth.    Ex- 
periments were  made  with  40  substances.    The  tabular 
summary  is  on  pp.  684-685.     Picric  acid  heads  the  list  for 
efficiency. 

(!8o).  REGNARD,  PAUL.  Influence  de  1'eau  oxygenee 
sur  la  fermentation.  Gaz.  med.  de  Paris 
T.  n,  6  ser.,  1880,  p.  358. 


('81).  CHAPPUIS,  E.  Action  de  1'ozone  sur  les  germes 
conitenus  dans  1'air.  Bull,  de  la  Soc.  chim. 
de  Paris,  I  sem.,  n.  s.,  T.  xxxv,  Paris,  1881, 
p.  290. 

('81).  BARNES,  J.  B.  The  antiseptic  properties  of 
cinnamic  acid.  Pharmaceut.  Jour,  and 
Transactions,  vol.  xn,  pp.  477-  478,  1881. 

('81).  JALAN  DE  LA  CROIX,  N.  Das  Verhalten  der 
Bakterien  des  Fleischwassers  gegen  einige 
Antiseptica.  Arch.  f.  exper.  pathol.  u. 
pharm.,  1881,  Bd.  xm,  pp.  175-255.  Ber.  d. 
•deutsch  chem.  Gesellsch.,  Bd.  xiv,  pp.  2,835- 
2,838. 

('81).  GOSSELIN,  L.,  ET  BERGERON,  A.  Recherches  sur 
la  valeur  antiseptique  de  certaines  substances 
et  en  particulier  de  la  solution  alcoolique  de 
Gaultheria.  Arch.  gen.  de  med.,  Paris,  1881, 
vol.  i,  VHI,  se.,  T.  7  (misprinted  6),  147  vol. 
de  la  collection,  pp.  16-29. 

('82).  BERT,  P.,  ET  REGNARD,  P.  Action  de  1'eau 
oxygenee  sur  les  matieres  organique  et  les 
fermentations.  C.  R.  des  se.  de  1'Acad.  des 
sci.,  T.  xciv,  1882,  pp.  1.383-1,386. 

('82).  BURCQ,  V.  Suir  Faction  desinfectante  et  anti- 
septique du  cuivre.  C.  R.  des  se.  de  Facad. 
des  sci.,  T.  xcv,  1882,  pp.  862-864. 

Workers  in  copper  escape  both  cholera  and  typhoid 
fever.  It  is  said  that  there  has  not  been  a  single  deatli 
from  either  disease  in  the  Societe  de  Bonaccord  (copper, 
bronze  and  brass  workers)  since  its  establishment  in  ;8ig. 

('82).  SCHIEFFERDECKER,  P.  Ueber  eine  neue  Injec- 
tionsmasse  zur  Conservirung  der  Leichen 
fiir  den  Praparirsaal.  Arch.  f.  Anat.  u. 
Entwickeltingsgesch.,  1882,  pp.  197-198. 

('82).  VULPIAN.  Etudes  experimentales  relatives  a 
Faction  que  peut  exercer  le  permanganate 
de  potasse  sur  les  venins,  les  virus  et  les 
maladies  zymotiques.  C.  R.  des  se.  de 
FAcad.  des  sci.,  T.  xciv,  1882,  pp.  613-617. 
Jour,  de  Pharm.  et  de  Chimie,  T.  vi,  5 
serie,  1882,  pp.  100-104. 

('86).  UNNA,  P.  G.  Ichthyol  und  Resorcin  als  Rep- 
rasentaten  der  Gruppe  reduzicrender  Heil- 
mittel.  Hamburg,  1886.  Unna's  Derma- 
tologische  Studien,  2  Heft,  pp.  1-85. 

('88).  NUTTALL,  GEO.  Experimente  iiber  die  bak- 
terienfeindlichen  Einfliisse  des  thierischen 
Korpers.  Zeitschr.  f.  Hyg.,  Bd.  iv,  1888,  pp. 
353-394,  i  plate. 

('88).  SALKOWSKI,  E.  Ueber  die  antiseptische  Wirk- 
ung  des  Chloroformvvassers.  Deutsche  med- 
ioin.  Wochenschrift,  1888,  Bd.  xiv,  pp.  309- 
3". 

('88).  LOEW,  O.  Physiologische  notizen  iiber  For- 
maldehyd.  Miinchen.  med.  Wochenschr., 
1888,  Bd.  xxxv,  pp.  412-413.  Physiol.  Ges. 
Miinchen  (1-6),  1888,  pp.  39-41.  Rev.  in 
Oh.  Centralb.,  1889,  LX  Jahrg.,  Bd.  i,  p.  90. 

C88).  BEHRING.  Ueber  QuecksilbersuWimat  in 
ei-weisshakigen  Fliissigkeiten.  Central!),  f. 
Bakt.,  1888,  Bd.  in,  pp.  27-30  and  64-66. 

('89).  FRAENKEL,  CARL.  Die  Ei-nwirkung  der  Kohlen- 
saure  auf  die  Lebensthiitigkcit  der  Mikro- 
organismen.  Zeitsch.  f.  Hyg.,  Bd.  v,  1889, 
PP-  332-362. 

('90).  ALTEHOEFER.  Ueber  die  Desinfectionskraft 
von  Wasserstoffsuperoxyd  auf  Wasser.  Cen- 
tralb.  f.  Bakt.,  1890,  Bd.  vm,  pp.  129-137. 

('90).  KIRCHNER,  M.  Untersuchungen  iiber  der  Ein- 
wirkung  des  Chloroforms  auf  die  Bakterien. 
Zeitschr.  f.  Hyg.,  Bd.  vm,  pp.  465-488,  1890. 


ANTISEPTICS   AND   GERMICIDES. 


251 


Coo).  SONNTAG,  HERMANN.  Ueber  die  Bedeutung 
des  Ozons  als  Desinficiens.  Zeitschr.  fur 
Hyg.,  1890,  Bd.  vm,  pp.  95-136. 

('91).  TIZZONI,  GUIDO,  y.  CATTANI,  G.  Ueber  die 
Widerstandsfahigkeit  der  Tetanusbacillen 
gegen  physikal-ische  und  chemische  Einwirk- 
ungen.  Archiv.  f.  exper.  Path.  u.  Pharm., 

1891,  Bd.  xxvin,  pp.  41-60. 

('91).  FROELICH,  O.     Ueber  das  Ozon,  dessen  Her- 
stellung  auf  elektrischem  Wege  und  dessen 
technische    Anwendungen,    inshesondere    in 
der       Gesundheitstechnik.         Gesundsheits- 
Ingenieur,  1891,  No.  16,  pp.  543-551. 
('91).  FROELICH,  O.     Ueber  das  Ozon,  dessen   Her- 
stellung  auf  eloktrischem  Wege  und  dessen 
technische  Anwendungen.    Electrotechnische 
Zeitschr.,  1891,  12  Jahrg.,  pp.  340-344. 
Contains  short  paragraph  on    physiological  action  of 
ozone.   Bacteria  living  in  water  are  killed  "  siimmtlich." 
No  experiments  with  pathogenic  bacteria.     It  is  still  a 
question  whether  bacteria  in  the  air  are  killed. 

CQI).  f.nti.Acii.  VAL.  Ueber  Lysol.  Zeitschr.  f. 
Hyg.,  Bd.  x,  1891,  pp.  167-196.  Also  a  sepa- 
rate. 

Lysol  is  more  active  than  carbolic  acid  or  creolin.  The 
hands  maybe  disinfected  in  a  i  per  cent,  solution  without 
soap.  Surgical  instruments  may  be  sterilized  in  \i  per 
cent  solution  without  the  least  injury.  Walls  may  bedis- 
infected  in  a  3  per  cent  solution.  It  is  to  man  tiie  least 
poisonous  of  the  antiseptics  of  its  class. 

('91).  FISCHER.     See  XL. 

('92).  SCHLUETER.     See  xxxvi. 

('92).  RICHTET,   CH.     De   Faction   de   quelques   sels 

metaliques  sur  la  fermentation  lactique.     C. 

R.  des  se.  de  1'Acad.  des  sci.,  T.  cxiv,  1892, 

pp.  1,494-1,496. 
f<;2).  OHLMUELLER.      Ueber    die     Einwirkung    des 

Ozons    auf    Bakterien.      Arbeiten    aus    dem 

Kaiserl.     Gesundheitsamte,    Bd.    vm,     1892, 

I  left  i,  pp.  229-251. 

Ozone  in  water  is  less  effective  as  a  germicide  in  pro- 
portion as  the  water  contains  more  and  more  dead 
organic  matter.  It  is  not  adapted  to  the  disinfection  of 
rooms. 

('92).  DELDRUECK.    See  xxxvi. 

('92).  HAMMER,  HANS.  Ueber  die  desinficirende 
Wirkung  der  Kresole  und  die  Herstellung 
netrtraler  wassriger  Kresollosungen.  II 
Mittheilung,  Arch.  f.  Hyg.,  Bd.  xiv,  1892, 
pp.  116-134. 

('92).  ARONSOHN,  HANS.  Ueber  die  antiseptischen 
Eigenschaften  des  Formaldehyde.  Berl.  klin. 
Wochenschr.,  1892,  Bd.  xxix,  No.  30,  pp. 
749-751- 

('92).  BERLIOZ,  F.,  AND  TRILLAT,  F.  Sur  les  pro- 
prieties des  vapeurs  du  formal  ou  aldehyde 
formique.  C.  R.  des  se.  de  1'Acad.  des  sci., 

1892,  T.  cxv,  pp.  290-292. 

('92).  HANKIN,  E.  L'action  bactericide  des  eaux  de 
la  Jumna  et  du  Gauge  sur  le  microbe  du 
cholera.  Ann.  de  1'Inst.  Pasteur,  T.  x,  pp. 
5"-523. 

Cholera  does  not  descend  the  rivers  in  India.  Bacteria 
are  much  rarer  in  these  rivers  than  in  European  rivers. 
The  filtered,  unboiled  river  water  has  a  decided  bacteri- 
cidal action  on  the  cholera  organism.  When  boiled  tilt- 
water  lost  its  germicidal  property. 

('93).  UE  CHRISTMAS.  J.  Sur  la  valeur  antiseptique 
de  1'ozone.  Ann.  de  1'Inst.  Pasteur,  T.  vn, 

1893,  PP-  776-78o. 
(*93)-  SCHII.D.    See  xvni. 
(*93)-  VOCES.    See  xxxvi. 


('93).  GREEN.  Ueber  den  Werth  der  Kupfersalze 
als  Desinfektionsmittel.  Zeitschr.  f.  Hyg., 
Bd.  xin,  1893,  pp.  495-511. 

States  that  copper  salts  have  considerable  value  as 
germicides,  especially  the  soluble  oiled.  Cuprum  bichlo- 
ratuni  is  considered  most  valuable.  This  is  the  only  cop- 
per salt  that  is  sufficiently  active  in  solutions  containing 
much  albumen.  For  the  treatment  of  wounds,  copper 
bichlorate  is  much  "belter  than  copper  sulphate. 

('93).  LOEW,  OSKAR.  Ein  natiirliches  System  der 
Giftwirkungen.  Mtinchen,  1893,  Wolff  und 
Liineburg,  pp.  vm,  136.  Rev.  in  Centralb.  f. 
Bakt.,  1893,  Bd.  xiv,  p.  234. 

('93).  GRUBER,  MAX.  Ueber  die  Loslichkek  der  Kre- 
sole in  Wasser  und  iiber  die  Verwendung 
ihrer  wassrigen  Losungen  zur  Desinfektion. 
Arch.  f.  Hyg.,  Bd.  xvn,  1893,  pp.  618-625. 

('94).  DIEUDONNE.     See  xxix. 

('94).  D'ARSONVAL  ET  CHARRIN.    See  xxxin. 

('94).  ABEL.    See  xvni. 

('94).  MIQUEL.  De  la  desinfection  des  poussieres 
seches  des  appartements;  and  Contribution 
nouvelle  a  1'etude  de  la  desinfection  par  les 
vapeurs  d'aldehyde  formique.  Ann.  de 
micr.,  T.  vi,  1894.  See  pages  257,  305,  396, 
520,  588,  and  621. 

('94).  POTTEVIN,  HENRI.  Recherches  sur  le  pouvoir 
antiseptique  de  1'aldehyde  formique.  Ann. 
de  1'Inst.  Pasteur,  T.  vm,  1894.  pp.  796-810. 

('94).  BOLTON,  MEADE.  The  effect  of  various  metals 
on  the  growth  of  certain  bacteria.  Internal. 
Med.  Mag.,  December,  1894,  PP-  812-822. 
Also  a  separate.  Reviewed  in  Am.  Nat., 
Oct.,  1895,  p.  933. 

('94).  SCHILOW,  P.  F.  Ueber  den  Einfluss  des  Was- 
serstoffstiperoxydes  auf  einige  pathogene 
Mikroorganismen.  St.  Petersb.  med.  Woch- 
enschr., 1894,  No.  6.  Rev.  in  Centralb.  f. 
Bakt.,  Bd.  xvi,  189.4,  PP-  42-43. 

Cholera  bacteria  were  destroyed  in  3  minutes  in 
1:200;  in  1:300  they  were  alive  after  I  hour.  Typhoid  ba- 
cilli, in  1:100  to  1:200,  were  killed  in  10  minutes  ;  in  1:1000, 
after  i  hour.  Anthrax  spores,  in  14  per  cent  solution  were 
killed  in  less  than  3  minutes;  a  2  per  cent  solution  killed 
them  in  less  than  one  hour;  a  l  per  cent  solution  did  not 
kill  in  I  hour.  Staphylpcoccus  pyogen.  aureus,  from  cul- 
tures i  day  old,  was  killed  in  1:100  in  less  than  lo  min- 
utes; in  1:200  it  required  more  than  15  minutes.  Diplo- 
coccus  pneumoniae  does  not  grow  in  bouillon  to  which 
hydrogen  peroxide  has  been  added  in  l:io,oooto  1:18,000. 
Solutions  of  1:200  destroyed  a  one-day  old  culture  in  15 
minutes. 

('94).  WALLICZEK,  HEINRICH.  Die  baktericiden 
Eigenschafiten  der  Gerbsaure  (Tannin  der 
Apotheken).  Centralb.  f.  Bakt.,  Bd.  xv, 
1894,  pp.  891-894. 

Tables  showing  effect  of  various  per  cents  of  tannin  on 
B.  coli,  B.  authracls,  and  Staphylococcus  aureus. 

('95).  BURCKHARD,  G.  Zwei  Beitrage  zur  Kenntnis 
der  Formalinwirkung.  Centralb.  f.  Bakt., 
xvni  Bd.,  1895,  pp.  257-264. 

Twenty  titles  are  cited  at  the  end  of  this  paper. 

('95).  VAN   ERMENCEM,   E.     De   la   sterilization   des 
eaux  par  1'ozone.     Ann.   de  1'Inst.   Pasteur, 
T.  ix,  1895,  pp.  673-709.     Rev.  in  Centralb. 
f.  Bakt.,  Bd.  xix,  1896,  pp.  836-838,  2  figs. 
Van  Ermengem's  report  is  favorable. 

('95).  SCHEPILEWSKY,  EUGEN.  Formaldeliyd  als 
Desinfektionsmittel.  (Dissert.)  St.  Peters- 
burg. 1895.  (Russisch.)  Rev.  in  Centralb. 
f.  Bakt.,  Bd.  xix,  1896,  pp.  794-796. 


252 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


('95).  GORIANSKY,  G.  J.  Sur  la  disinfection  des 
crachats  phtisiques  et  des  cultures  tuber- 
culeuses  par  les  solutions  alcalines  de  good- 
ran  et  de  vwiaigre  de  bois.  Arch,  des^  sci. 
biol.,  pub.  par  1'Inst.  imp.  de  med.  exp.  a  St. 
Petersburg,  Tome  3,  1895,  pp.  148-166. 

Wood-vinegar  is  a  very  energetic  disinfectant.  lu  quan- 
tity equal  to  the  volume  of  sputum,  and  acting  for  6 
hours,  it  was  found  entirely  efficient.  Exposure  of  4 
hours  is  not  sufficient  in  some  cases  to  destroy  B.  tuber- 
culosis in  sputum.  In  pure  culture  the  organism  is  de- 
stroyed by  exposure  for  l  hour  to  this  acid. 

('95).  D'ARSONVAL.  Sur  la  production  de  1'ozone 
concentre  et  sur  ses  effets  bactericides.  C. 
R.  des  se.  et  mem.  de  la  soc.  de  foiol.,  Paris, 
10  se.,  T.  II,  1895,  pp.  500-502. 

The  writer's  experiments  were  negative,  and  he  is  very 
skeptical  as  to  germicidal  power  of  ozone. 

('95).  FISCHER.    See  XL. 

('96).  WALTER,  K.     Zur  Bedeutung  des  Formalins, 

bezw.       Formaldehyde     als     Desinfekrions- 

mittel.    Zeitschr.  f.  Hyg.,  Bd.  xxi,  1896,  pp. 

421-451.    Rev.  in  Cerrtralb.  f.  Bakt.,  xx  Bd., 

1896,  p.  280. 
('97).  IWANOFP,  W.   A.     Zur  Frage  iiber   das  Ein- 

dringen  der  Formalindampfe  in  die  organ- 

ischen    Gewebe.      Centralb.    f.    Bakt.,    xxn 

Bd.,  1897,  pp.  50-58. 

Formalin  vapor  did  not  penetrate  rapidly  into  the 
depths  of  the  tissues  tested  (livers  of  rabbits  and  guinea 
pigs). 

('97).  WEYLAND,  J.  Desinfektionswirkung  und 
Eiweissfallung  chemischer  Korper.  Cen- 
tralb. f.  Bakt.,  xxi  Bd.,  1897,  pp.  798-802. 

('97).  FuERBRlNGER  UND  PREYHAN.  Neue  Unter- 
suchungen  ii'ber  die  Desinfektion  der  Hande. 
Deutsche  med.  Wochenschr.,  1897,  No.  6. 
Rev.  in  'Centralb.  f.  Bakt.,  xxi  Bd.,  1897,  pp. 
708-710. 

Authors  recommend  alcohol.  A  2  per  cent  solution  of 
mercuric  chloride  is  still  better.  Both  may  be  used,  the 
latter  preceded  by  the  former. 

('97).  PODGORNY,  K.  M.  Effect  of  iodine  on  patho- 
genic bacteria.  Thesis  of  St.  Petersburg, 
No.  36,  1897,  pp.  74. 

('97).  SCHUMBURG.  Ein  neues  Verfalhren  zur  Her- 
stellung  keimfreien  Trinkwassers.  Deutsche 
med.  Wochenschr.,  Bd.  xxm,  1897,  No.  10, 
pp.  145-146. 

Bromide  treatment. 

('98).  MINERVINI,  RAFAEL.  Ueber  die  baktericide 
Wirkung  des  Alkohols.  Zeitschr.  f.  Hyg., 
Bd.  xxix,  1898,  pp.  117-148.  Bibliography 
of  18  titles. 

Ethyl  alcohol  has  only  a  weak  bactericidal  action.  It 
is  most  active  in  concentrations  of  50  to  7o  per  cent.  Al- 
coholic solutions  of  antiseptic  substances  are  less  active 
than  water  solutions. 

('08).  SCHULTZ,  N.  De  I'actipn  des  antiseptiques  sur 
•le  bac.  pestis  hommis  et  de  la  desinfection 
d'effeots  et  de  locaux  contamines  par  la  peste 
bubonique.  Arch,  des  >sci.  biol.  publiees  par 
1'inst.  imper.  de  ined.  exper.  a  St.  Peters- 
bourg,  T.  vi,  1808,  pp.  397-426,  i  plate. 

The  appearance  of  the  bacteria  subjected  to  the  anti- 
septics, as  shown  on  the  plate,  strongly  suggests  the  ap- 
pearance of  organisms  in  old  cultures,  viz.:  involution 
forms,  and  the  two  phenomena  may  be  due  to  the  same 
cause,  the  involution  forms  arising'from  the  harmful  ac- 
tion of  products  excreted  by  the  bacteria,  or  arising  from 
the  action  of  siibstances  developed  in  the  media  as  the 
result  of  bacterial  occupation. 


('98).  Fi.uEGGE,  C.  Die  Wohnungsdesinfektion  durch 
Formaldehyd.  Zeitschr.  f.  Hyg.,  Bd.  xxix, 
1898,  pp.  276-308. 

('98).  FERMI.    See  xxxvi. 

('98).  POPOFF,  S.  P.  Vergleichende  Studien  iiber  die 
desinfizierende  Wirkung  reiner  Sublimat- 
losungen  und  Kombinationen  derselben  mit 
anderen  Desinficientien.  (Diss.)  St.  Peters- 
burg, 1898.  Rev.  in  Centralb.  f.  Bakt.,  xxv 
Bd.,  1899,  pp.  331-332. 

The  power  of  i  per  cent  sol.  mercuric  chloride  is  in- 
creased very  decidedly  by  addition  of  i  per  cent  hydro- 
chloric acid,  or  i  per  cent  phenol,  and  by  the  addition 
of  2  per  cent  phenol  a  still  more  effective  mixture  is 
obtained.  Addition  of  i  to  2  per  cent  NaCl  weakened  the 
effect  of  the  i  per  cent  sublimate  solution  on  some  or- 
ganisms, but  increased  it  on  others. 

('98).  LUCAS-CHAMPIONNJERE.  Sur  la  valeur  anti- 
septique  de  1'eau  oxygenee.  Bull,  de  I'acad. 
de  med.,  1898,  T.  xi,,  serie  3,  Paris,  pp.  599- 

6i7. 

('99).  MARMIER  ET  ABRAHAM.  La  sterilisation  indus- 
trielle  des  eaux  potables  par  1'ozone.  Rev. 
d'hyg.  et  de  Police  Sanitaire,  Paris,  1899,  T. 
xxi,  pp.  540-554. 

Great  things  are  claimed  for  this  method.  Only  some 
specimens  of  Bacillus  subtilis  are  said  to  have  escaped 
destruction,  and  of  these  only  one  individual  for  each 
15  cc.  of  water  treated  with  a  concentration  of  ozone 
equal  to  6  milligrams  per  litre  of  air. 

('99).  STABLER,  EDUARD.  Ueber  die  Einwirkung  von 
Kochsalz  auf  Bakterien,  die  bei  den  soge- 
nannten  Fleischvergiftungen  eine  Rolle 
spielen.  Arch.  f.  Hyg.,  Bd.  xxxv,  1899,  pp. 
40-82.  Rev.  in  Centralb.  f.  Bakt.,  xxvi  Bd., 
1899,  p.  411. 


2  to  3  clays  tney  were  injured,  nut  alter  tnai  iney  maae 
a  luxuriant  growth.  The  toleration  limit  for  B.  coli  and 
B.  enteritidis  is  between  7  and  8  per  cent  of  NaCl,  and  that 
of  B.  morbificans  bovis  between  8  and  10  per  cent.  The 
writerof  this  abstract  found  certain  plant  bacteria  much 
more  sensitive  to  salt,  e.  g.  Ps.  hyacinthi  was  restrained 
by  1.5  per  cent. 

('99).  BLISS,  C.  L.,  AND  NOVY,  PR.  G.  Action  of 
formaldehyde  on  enzymes  and  on  certain 
proteids.  The  Jour.  Exp.  Medicine,  vol.  iv, 
1899,  pp.  47-80. 

('99).  WEYL,  TH.  Keimfreies  Trinkwasser  mittels 
Ozon.  Centralb.  f.  Bakt.,  xxvi  Bd.,  1899, 
<PP-  15-32,  with  i  fig. 

"Ozone  is  a  specific  bacterial  poison."  It  is  recom- 
mended for  sterilizing  drinking  water. 

('99).  KOCH,  E.,  AND  FUCHS,  G.  Ueber  den  anti- 
bakteriellen  Wert  des  Acrolein.  Centralb. 
f.  Bakt.,  xxvi  Bd.,  1899,  pp.  560-563. 

Acrolein  is  a  substance  related  to  formaldehyd.  In 
0.25  to  0.5  per  cent  solutions  it  proved  more  effective  than 
formaldehyd  on  a  number  of  uon-sporiferous  organisms. 

('99).  CALMETTE,  A.  Rapport  sur  la  sterilization  in- 
dustrielle  des  eaux  potables  par  1'ozone. 
Ann.  de  I'lnst.  Pasteur,  1899,  T.  xm,  pp. 
344-357- 

A  favorable  report  on  sterilization  of  water  by  ozone 
by  a  committee,  of  which  Calmette  was  secretary.  They 
recommended  the  system  of  Marmier  and  Abraham  for 
the  city  of  Lille. 

('99).  Roux  ET  CALMETTE.  Sterilization  of  water  by 
ozone.  Rapport  presente  a  la  miunicipalite 
de  Lille,  fevrier,  1899. 

Not  seen. 


ANTISEPTICS   AND    GERMICIDES;    CHEMOTROPISM,    ETC. 


253 


('99).  MARPMANN.  Die  baktericide  Wirkung  des 
Fluornatriunis  und  der  Nachweis  dessolben 
in  Nahruiiifiiiitteln.  Central!),  f.  Bakt.,  Bd. 
xxv,  1899,  pp.  309-311. 

Considers  sodium  fluoride  a  valuable  disinfectant  for 
all  bacteria  in  nutrient  media.  Sodium  fluoride  is  said 
to  be  scarcely  more  harmful  to  man  tliau  common  salt. 
In  beer  wort  in  doses  of  i  gram  per  litre  it  inhibits  the 
growth  of  bacteria  without  lessening  that  of  the  yeast. 

('oo).  OTTO.  Sterilization  of  water  by  ozone.  Bulle- 
tin de  la  Spciete  des  ingenieurs  civils  de 
France,  fevrier,  1000. 

('01).  MAYER,  EUGEN,  UND  WOLPERT,  HEINRICH. 
Beitriige  zur  Wohungsdesinfektion  durch 
Formaldehyd :  I.  Die  zweckmassigste  Form 
des  Verdampfungsapparats.  n.  Einfluss  der 
Temperatur  auf  die  Desinfektipnswirkung. 
in.  Verstarkung  der  Desinfektionswirkung 
durch  kiinstliche  Luftmischung  (Vorl. 
Mitt.).  Hyg.  Rdsch.,  Berlin,  xi  Jahrg.,  1901, 
PP.  IS3-I58. 

('01).  SMITH,  ERWIN  F.  Growth  of  bacteria  in  the 
presence  of  chloroform  and  -thymol.  Science, 
n.  s.,  vol.  xm,  p.  327,  March  i,  1901.  See 
also  Jour.  Boston  Soc.  Med.  Sci.,  vol.  v,  p. 
375,  and  Central!),  f.  Bakt.,  1901,  i  Abt.,  Bd. 
xxix,  pp.  445-446. 

('oi).  HESS,  OTTO.  Der  Formaldehyd.  Seine  Daj- 
stellung,  Eigenschaften,  und  seine  Ver- 
wendung  als  Konservierimgs-therapeutisches 
und  Desinfektionsmittel  unit  besonderer 
Beriicksichtigung  der  Wohnungsdesinfec- 
tion.  Marburg,  N.  G.  Elwert,  2  Aufl.,  1901, 
pp.  iv,  129. 

('02).  ROLLY.  Zur  Analyse  der  Borax- und  Borsaure- 
wirkung  bei  Faulnissvorgangen,  nefest 
Studien  iiber  Alkali-  und  Saureproduktion 
der  Faulnissbakterien.  Arch.  f.  Hyg.,  Bd. 
xu,  1902,  pp.  348-4°5- 

('02).  PRESCOTT,  SAMUEL  C.  Antiseptics  and  their 
use  in  the  preservation  of  food.  Technology 
Quarterly.  Vol.  xv,  1902,  pp.  335-342. 

(*02).    HlLL,    HlBBERT    W.,     AND    RlCKARDS,    BuRT    R. 

Notes  on  formaldehyd.  Proc.  Am.  Pub. 
Health  Asso.,  3oth  Ann.  Meeting,  New  Or- 
leans, Dec.,  1902.  Also  a  separate,  pp.  12. 

('02).  SCHUEDER.  Entgegnung  auf  die  Schumburg' 
sche  Arbeit:  "Das  Wasserreinigungsver- 
fahren  mit  Brom"  und  die  Arbeit  von  A. 
Pfuhl :  "Zu  den  Schiider'schen  Prufungsver- 
suchen  des  Bromverfahrens  nach  Schum- 
burg." Zeitsch.  f.  Hyg.,  Bd.  xxxix,  1902, 
PP-  532-539- 

('02).  SCHUMBURG,  WILHELM.  Zu  der  Schiider'schen 
Entgegnung  beziiglich  des  Bromverfahrens 
zur  Trinkwasser-Reinigung.  Zeitsch.  f. 
Hyg.,  Bd.  XL,  1002,  pp.  199-202. 

('02).  GREEN,  A.  B.  The  disinfectant  action  of 
chloroform  and  various  other  substances  on 
the  specific  and  extraneous  micro-organisms 
of  vaccine.  Rep.  Med.  Off.  Loc.  Gov.,  Lon- 
don, 1902,  p.  639-663. 

('02).  ENGELS,  EUGEN.  Das  Schumburg'sche  Ver- 
fuhren  der  Trinkwasserreinigung  mittels 
Brom.  Central!),  f.  Bakt.,  Abt.  i,  Bd.  xxxi, 
Originale,  1902,  pp.  651-670. 

("02).  KONRADI,    DANIEL.     Ueber     die     baktericide 
Wirkung   der   Seifen.     Arch.    f.    Hyg.,   Bd. 
XLIV,  1902,  pp.  101-112. 
Recommends  a  resorcin  soap. 


('02).  MAYER,  EUGEN,  UND  WOLPERT,  HEINRICH. 
Ueber  die  Verfa'hren  und  Apparate  zur 
Entwicklung  von  Formaldehyd  fitr  die 
Zwecke  der  Wohnungsdesinfektion.  Arch, 
f.  Hyg.,  Bd.  XLIII,  1902,  pp.  157-169. 

("02).  COHN,  ERNST.  Ueber  den  antiseptischen  Wert 
des  Argentum  colloidale  Crede  und  seine 
Wirkung  bei  Infektion.  Diss.  Konigsberg  i. 
Pr.  (Druck  v.  M.  Hiller),  1902,  p.  57. 

('03).  RICKARDS,  BURT  RANSOM.  A  comparison  of 
some  of  the  more  common  liquid  disin- 
fectants. Jour.  Mass.  Asso.  of  Boards  of 
Health,  vol.  xni,  No.  3,  Oct.,  1903,  pp.  70-76. 

('03).  FREER,  PAUL  C.,  AND  NOVY,  FREDR.  G.  On  the 
organic  peroxides.  Vaughan  Quarter  Cen- 
tury Book,  pp.  63-127.  Ann  Arbor,  1903. 

"  Acetyl  and  benzoyl  hydrogen  peroxides  are  extremely 
germicide!,  and  easily  rank  with  the  most  active  disin- 
fectants. *  *  Hydrogen  peroxide  is  considerably 
weaker  than  these  organic  peroxides.  The  activity  of 
the  peracids  and  of  hydrogen  peroxide  is  not  due  to 
active  oxygen,  but  is  probably  due  to  the  acid  ions." 

('03).  RIDEAL,  SAMUEL.  Disinfection  and  the  preser- 
vation of  food,  together  with  an  account  of 
the  chemical  substances  used  as  antiseptics 
and  preservatives.  3d  ed.,  London,  Sanitary 
Pub.  Co.,  Ltd.;  New  York,  John  Wiley  & 
Sons,  1903,  pp.  494. 

('04).  KONRADI,    DANIEL.     Weitere    Untersuchungen 
ueber  die  bakterizide  Wirkung  der  Seifen. 
Centralb.  f.  Bakt.,  i  Abt.,  Originale,  xxxvi 
Bd.,  1904,  pp.  151-160. 
St.  Laceleau  soap  is  actively  bactericidal, 

('05).  KRAEMER,  HENRY.  The  Oligodynamic  Action  of 
Copper  Foil  on  Certain  Intestinal  Organisms. 
Proc.  Am.  Phil.  Soc.,  vol.  XLIX,  pp.  51-65. 
Phila.,  1905.  Also  a  separate. 


XXXIX.    Chemotropism,  Thermotropism,  Geotrop- 
ism,  Contact-Irritation,  Etc. 

PFEFFER,     W.       Lokomotorische     Richtungs- 
bewegungen  durch  chemisdhe  Reize.    Unter- 
suchungen aus  dem  bat.  Institut  Tiibingen, 
i  Bd.,  1884,  pp.  363-482. 
The  chapter  on  Spaltpilze  begins  on  p.  449. 

DUBOIS,  R.  Influence  du  magnetisme  sur 
l'orien<tation  des  colonies  microbiennes.  C. 
R.  des  se.  et  inem.  de  la  soc.  de  biol.,  Paris, 
1886,  8  se.,  T.  in,  pp.  127-128. 

PFEFFER,  W.  Ueber  chemotaktische  Bewe- 
gungen  von  Bakterien,  Flagellaten  und  Vol- 
vocineen.  Untersuch.  a.  d.  bot.  Inst.  zu 
Tubingen,  1888,  Bd.  n,  Heft  3,  pp.  582-661. 

ALI-COHEN,  CH.  H.  Die  Chemotaxis  als  Hiilfs- 
mittel  der  bakteriologischen  Forschung. 
Centralb.  f.  Bakt.,  vin  Bd.,  1890,  pp.  161-167. 

BOYCE  AND  EVANS.  Upon  the  action  of  gravity 
on  Bacterium  Zopfii.  Communication  made 
to  the  Royal  Society,  Feb.,  1893.  Rev.  in 
Centralb.  f.  Bakt.,  Bd.  xv,  1894,  pp.  568-569. 

ROTH,  A.  Ueber  das  Verhalten  beweglicher 
Mikroorganismen  in  stromenden  Fliissig- 
keiten.  Deutsch.  med.  Wochenschr.,  1893, 
No.  15,  PP.  351-352- 

In  streaming  fluids  this  author  observed  in  motile  bac- 
teria a  decided  tendency  to  move  against  the  current. 

('94).  MIYOSHI,    MAN  ABU.     Ueber    Chemotropismus 
der   Pilze.     Botanische   Zeitung,    1894,   Hft. 
i,  col.  1-28,  with  i  table. 
Deals  only  with  fungi. 


('84). 


('86). 


('88). 


('{jo). 


('93). 


('93). 


254 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('94).  BEYERINCK,  M.  W.  Ueber  Thermotaxis  bei 
Bakterium  Zopfii.  Centralb.  f.  Bakt.,  Bd. 
xv,  1894,  p.  799. 

Refers  to  thermotaxis  the  movements  of  the  threads 
of  Bact.  Zopfii,  which  Boyce  and  Evans  supposed  to  be 
due  to  geotropism. 

('or).  JENNINGS,  H.  S.,  AND  CROSBY,  J.  H.  Studies 
on  reactions  'to  stimuli  in  unicellular  or- 
ganisms, vn.  The  manner  in  which  bac- 
teria react  to  stimuli,  especially  to  chemical 
stimuli.  Am.  Jour,  of  Physiol.,  vol.  vi,  1901, 
PP-  3J-37-  Also  a  separate. 

The  movement  of  unicellular  organisms  toward  or 
away  from  chemical  substances  is  said  to  be  due  to  a 
"motor  reflex  "  comparable  to  that  of  the  ciliate  infu- 
soria, aud  not  to  chernotaxis,  as  stated  by  Pfefier  and 
others.  These  conclusions  agree  with  those  of  Rothert. 

('oi).  ROTHERT,  W.  Beobachtungen  und  Betrach- 
tungen  fiber  tactische  Reizerscheinungen. 
Flora  oder  Allgemeine  Botanische  Zeitung, 
88  Bd.,  1901,  Hft.  in,  pp.  371-421. 

('02).  CLARK,  JUDSON  F.    On  the  toxic  properties  of 
some  copper  compounds  with  special  refer- 
ence to   Bordeaux  mixture.     Botanical   Ga- 
zette, vol.  xxxni,  1902,  pp.  26-48,  7  figs. 
Controverts  Miyoshi  on  chemolrop  am. 

('03).  ROTHERT,  W.  Ueber  die  Wirkung  des  Aethers 
und  Chloroforms  auf  die  Reizbewegungen 
der  Mikroorganismen.  Jahrb.  f.  Wiss.  Bot, 
Bd.  xxxix,  1903,  pp.  1-70. 

The  variability  in  sensitiveness  of  the  same  organism 
at  different  times  was  most  disturbing.  "  Material 
which  to-day  is  strikingly  chemotactic  or  phototactic, 
may  be  to-morrow  unusable.  Especially  striking  and 
perplexing  was  the  behavior  of  Spirillum  undula,  this 
classical  object  for  chernotaxis  and  osmotaxis,  with 
which  Pfeffer  made  his  celebrated  investigations,  and 
with  which  in  former  years  I  confirmed  the  experiments 
of  Pfeffer." 


XL.     Osmotic  Pressures. 

CQI).  WLADIMIROFF,  ALEXANDER.  Osmotische  Ver- 
suohe  an  lebenden  Bakterien.  Zeitschr.  f. 
physikalische  Chem.,  Bd.  vn,  1891,  pp.  529- 
543- 

('91).  WLADIMIROFF,  ALEXANDER.  Biologische  Studien 
an  Bakterien.  i.  Uber  das  Verhalten 
beweglicher  Bakterien  in  Losungen  von 
Neutralsalzen.  Zeitschr.  f.  Hyg.,  1891,  Bd. 
x,  pp.  89-110. 

('91).  FISCHER,  ALFRED.  Die  Plasmolyse  der  Bak- 
terien. Ber.  iiber  die  Verhandlungen  d.  K. 
sachs.  Ges.  d.  Wissenschaften.  Mathetn.- 
phys.  Classe,  Leipzig,  1891,  Bd.  XLIII,  pp. 
52-74,  i  plate. 

Plasmolysis  was  either  definitely  established  or  ren- 
dered extremely  presumptive  (smaller  forms)  for  I7kinds 
of  bacteria.  The  concentration  which  induced  it  varied 
in  most  cases  from  0.5  to  5  per  cent.  In  a  few  cases  the 
least  concentration  which  would  induce  plasmolysis  was 
not  determined.  Generally  strengths  of  i  to  2  percent 
sodium  chloride  were  sufficient.  "  Die  untere  Grenze 
liegt  fast  fiir  alle  Bacterien  bei  I  per  cent  oder  0.75  per 
cent  NaCl." 

('95).  FISCHER,  ALFRED.  Neue  Beobachtungen  iiber 
Plasmolyse  der  Bakterien  in  Untersuchungen 
iiber  Bakterien.  Jahrb.  f.  wissensch.  Bot., 
Berlin,  1895,  Bd.  xxvn,  pp.  1-34. 

Coi).  D'ARSONVAL.    See  xxxni. 


XLI.     Chemical  Analysis  of  Bacteria. 

('79).  NENCKI,  M.,  UND  SCHAFFER,  F.  Ueber  die 
ohemische  Zusammensetzung  der  Faulniss- 
bakterien.  Jour.  f.  Praktische  Chemie,  neue 
Folge,  Bd.  xx,  1879,  pp.  443-466,  i  fig.,  i 
plate.  Also  a  separate. 

('81).  SCHAFFER,  F.  Zur  kenntniss  des  Mykoproteins. 
Journal  f.  Prak.  Chemie,  neue  Folge,  Bd. 
xxill,  1881,  pp.  302-304. 

('86).  BROWN,  ADRIAN  J.  On  an  acetic  ferment  which 
forms  cellulose.  Journal  Chem.  Soc.  Trans., 
London,  vol.  XLIX,  pp.  432-439. 

('87).  BROWN,  ADRIAN  J.  Note  on  the  cellulose 
formed  by  Bacterium  xylinum.  Journ. 
Chem.  Soc.,  London,  Trans.,  vol.  LI,  1887, 
p.  643. 

('87).  VINCENZI,  LIVIO.  Uebor  die  chemischen  Be- 
standteile  der  Spaltpilze.  Zeitschr.  f. 
physiolog.  Chemie,  1887,  Bd.  xi,  pp.  181-183. 

('88).  HAMMERSCHLAG.  Ueber  bacteriologisch- 
chemische  Untersuchung  der  Tuberkel- 
bacillen.  Verhandlungen  der  Schweizeri- 
schen  Naturf.  Gesselsch.  in  Solathurn,  Au- 
gust, 1888,  71  Jahresversammlung,  pp.  85-86. 

CP3)-  CRAMER,  E.  Die  Zusammensetzung  der  Bak- 
terien in  ihrer  Abhangigkeit  von  dem  Nahr- 
rnaterial.  Arch.  f.  Hyg.,  Bd.  xvi,  Heft  2, 
1893,  pp.  151-195. 

('93).  NISHIMURA,  TOYOSAKU.  Untersuchung  fiber 
die  chemische  Zusammensetzung  eines 
Wasserbacillus.  Arch.  f.  Hyg.,  Bd.  xvm, 
1893,  PP.  318-333. 

('93).  DREYFUSS,  ISIDOR.  Ueber  das  Vorkommen 
von  Cellulose  in  Bacillen,  Schi.mmel-  und 
anderen  Pilzen.  Zeitschr.  f.  physiol.  chem., 
Bd.  xvm,  1893,  pp.  358-379- 

The  conclusion  is  that  cellulose  occurs  in  hay  bacilli 
and  in  pus  bacilli. 

('95).  CRAMER,  E.  Die  Zusammensetzung  der 
Cholerabacillen.  Arch.  f.  Hyg.,  Bd.  xxn, 
1895,  PP-  167-190. 

('98).    DE    SCHWEINITZ,    E.    A.,    AND    DORSET,    MARION. 

The  mineral  constituents  of  the  tubercle 
bacilli.  Centralb.  f.  Bakt.,  Bd.  xxni,  1898, 
PP.  993-995- 

('02).  KRESLING,  K.  I.  De  la  substance  grasse  des 
bacilles  de  la  tuberculose.  Arch,  des  sci. 
biol.,  publiees  par  1'inst.  imp.  de  med.  exper. 
a  St.  Petersbourg,  T.  ix,  1902,  pp.  359-376. 


XLI  I.     Distribution  of  Bacteria— Geographical  and 
Attitudinal. 

(Deserts,  mountains,  arctic  regions,  sea  air,  depths 
of  the  sea,  deep  wells,  surface  soils,  air  at  the  earth's 
surface,  and  at  higher  levels.) 

('81).  MIQUEL,  P.  Sur  le  dosage  des  bacteries  dans 
les  poussieres  et  dans  le  sol.  Bull.  Soc.  Bot. 
de  France,  1881,  T.  xxvin,  ser.  3,  pp.  44-51. 

('82).  TYNDALL,  JOHN.  Essays  on  the  floating-mat- 
ter of  the  air  in  relation  to  putrefaction  and 
infection.  New  York,  D.  Appleton  &  Co., 
1882,  pp.  xix,  338. 

('83).  MIQUEL,  P.  Les  organismes  vivants  de  1'at- 
mosphere,  pp.  vm,  310.  Paris.  1883.  Gau- 
thier-Villars. 


DISTRIBUTION    OF    BACTERIA — GEOGRAPHICAL    AND    ALTITUDINAL. 


255 


('83).  MNJUEL,  P.  Nouvelles  recherches  sur  les  bac- 
teries  atmospheriques  effectuees  a  1'observa- 
toire  de  Montsouris.  Ann.  de  1'observatoire 
de  Montsouris  pour  1'an  1883,  pp.  391-437. 
('83).  MIQUEL,  P.  De  la  purete  en  microbes  de  1'air 
des  montagnes  et  de  quelques  districts  de  la 
Suisse.  La  semaine  medicale,  1883,  pp.  274- 
276. 

('83).  GIACOSA,  PIERO.  Studii  sui  corpuscoli  organ- 
izzati  dell'aria  sulle  alte  montagne.  Atti  R. 
Accad.  d.  sci.  di  Torino,  vol.  xviil,  pp.  263- 
272,  1883,  I  plate. 

('84).  DE  FREUDENREICH,  ED.  Des  microbes  de  1'air 
des  montagnes.  La  semaine  medicale,  11 
septembre,  1884,  pp.  361-362. 

Relates  to  scarcity  of  bacteria  in  the  air  at  high  alti- 
tudes. In  the  summer,  in  the  Beruese  Alps,  in  four  places 
at  altitudes  varying  from  2.000  to  4,000  meters,  a  total  of  2700 
litres  of  air  were  aspirated  without  finding  any  bacteria. 
The  following  summer  tests  were  made  on  a  glacier  2,900 
meters  high  and  on  the  summit  of  a  mountain  2,366 
meters  high,  vegetation  reaching  nearly  to  the  top.  On 
the  glacier  2,000  litres  of  air  were  aspirated  in  six  por- 
tions. Two  of  the  sowings  remained  sterile.  One  gave  a 
micrococcus,  another  B.  subtilis,  one  after  more  than  15 
days  gave  a  torula.  and  one  a  mold.  "  II  nous  resterait  2 
bacteries  pour  2,000  litres,  soit  un  par  metre  cube."  In 
Bern  the  author  says  he  obtains  hundreds  and  thousands 
of  bacteria  per  cubic  meter  using  the  same  delicate 
methods. 

The  experiments  on  the  mountain  yielded  8  bacteria 
from  2,325  litres  of  air. 

('84).  DE  FREUDENREICH,  EDOUARD.  Recherches  sur 
les  organismes  vivants  de  1'air  des  hautes 
altitudes.  Archiv.  des  sci.  physiques  et  nat- 
urelles,  3e  Periode,  T.  douzieme,  Geneve, 
1884,  pp.  365-387. 

Author  shows  that  the  purity  of  the  air  on  mountains 
is  much  greater  than  former  writers  have  supposed. 

('84).  MIQUEL,  P.  Des  organismes  microscopique  de 
1'air  de  la  mer.  La  semaine  medicale,  1884, 
pp.  90-92. 

('84).  HESSE,  W.  Ueber  quantitative  Bestimmung 
der  in  der  Luft  Enthaltenen  Mikrporgan- 
ismen.  Mitfrh.  a.  d.  K.  Gesundheitsajnte, 
Berlin,  vol.  n,  1884,  pp.  182-207. 
('85).  MIQUEL,  P.  Septieme  memoire  sur  les  or- 
ganismes microscopiques  de  1'air  et  des  eaux. 
Annuaire  de  1'observatoire  de  Montsouris 
pour  Fan  1885,  pp.  467-611. 

The  part  relating  to  microorganisms  in  the  sea-air 
begins  on  p.  514.  This  subject  is  also  treated  inthereport 
of  the  observatory  for  1886,  pp.  535-550. 

('86).  BEUMER. .     Zur     Bakteriologie     des     Bodens. 

Deutsche  med.   Wochenschr.,  Bd.   xn,   1886, 

pp.  464-466. 
('86).  FISCHER,  B.    Bakteriologische  Untersuchungen 

auf  einer  Reise  nach  Westindien.     Zeitschr. 

f.  Hyg.,  Bd.  I,  1886,  pp.  421-464. 

This  paper  discusses  the  microorganisms  and  spores 
found  in  sea -air. 

('86).  ADAMETZ,  LEOPOLD.  Untersuchungen  iiber  die 
niederen  Pilze  der  Ackerkrume.  Inaugural 
Dissertation,  78  pp.,  2  Taf.,  Leipzig,  1886. 
Rev.  in  Centralb.  f.  Bakt,  1887,  Bd.  I,  pp. 
8-10. 

('87).  PETRI,  R.  J.  Zusammenfassender  Bericht  iiber 
Nachweis  und  Bestimmung  der  pflanzlichen 
Microorganismen  in  der  Luft.  Centralb.  f. 
Bakt.,  Bd.  n,  1887,  pp.  113-118  und  151-158. 


('87).  MAGGIORA,  A.  Ricerche  quantitative  sui  micro- 
organism! del  suolo  con  speciale  riguardo, 
all'inquinazione  del  medesimo.  Giornale 
della  R.  Accademia  de  medicina  di  Torino, 
1887,  vol.  xxxv.  Series  3,  pp.  153-172. 

('87).  FRAENKEL,  CARL.  Untersuchungen  iiber  das 
Vorkommen  von  Mikroorganismen  in  ver- 
schiedenen  Bodenschichten.  Zeitschr.  f. 
Hyg.,  1887,  Bd.  u,  pp.  521-582. 

('89).  REIMERS,  JOHN.  Ueber  den  Gehalt  des  Bodens 
an  Bacterien.  Zeitschr.  f.  Hyg.,  1889,  Bd. 
vn,  pp.  307-346. 

('89).  REIMERS,  JOHN.  Ueber  den  Gehalt  des  Bodens 
an  Bacterien.  Inaug.  Dissert.  Jena,  1889, 
8vo.,  44  pp.,  Leipzig,  Veit  u.  Comp.  Re- 
viewed in  Centralb.  f.  Bakt.,  I  Abt,  Bd.  x, 
1891,  p.  489. 

"  Die  Zone  dieser  plotzlichen  Keimverminderung  liegt 
im  Jeneuser  Boden — wie  im  Berliner — zwischen  i  und  2 
metres." 

('90).  KRAMER.    See  in. 
('91).  MANFREDI.    See  XLIII. 

('93).  CRISTIANI,  H.  Analyse  bacteriologique  de  1'air 
des  hauteurs  puise  pendant  un  voyage  en 
ballon.  Ann.  de  1'Inst.  Pasteur,  T.  vn  pp 
665-671. 

At  elevations  above  the  soil  of  i.oco  meters  and  upward 
the  cultures  remained  sterile  although  iu  each  case  10 
litres  of  air  was  allowed  to  bubble  through  the  culture 
media.  Even  at  much  lower  levels  the  majority  of 
colonies  are  believed  to  have  come  from  the  earth  indi- 
rectly, i.  e.,  by  way  of  the  balloon. 

('93).  DUCLAUX,  E.  La  distribution  de  la  matiere 
organique  et  des  microbes  dans  le  sol.  Revue 
critique.  Ann.  de  1'Inst.  Paseur,  T.  vn, 
1893,  pp.  823-833. 

('97).  MORITZ,  Orro,  UND  NEUMANN,  R.  O.  Ueber 
einige  bakteriologische  Wasseruntersuchun- 
gen  im  Atlantischen  Ozean.  Centralb.  f. 
Bakt.  2  Abt,  Bd.  xin,  1904,  pp.  481-489. 

("99).  LEVIN.  Les  microbes  dans  les  regions  arc- 
tiques.  Ann.  de  1'Inst.  Pasteur,  T.  xm,  1899, 
PP.  558-567. 


The  arctic  air  was  tested  in  twenty  places,  approxim- 
ately the  same  amount  being  filtered  at  each  place.  A 
total  of  21, 6po  litres  of  this  filtered  air  yielded  three  bac- 
terial colonies  and  a  few  mold  spores.  The  surface  waters 
of  the  arctic  also  contain  few  bacteria.  Ninety  samples 
of  water  were  taken  from  thesea  at  greatdepths  (i.ooo  to 
3,000  metres).  These  samples  also  contained  bacteria  of 
several  kinds  but  in  small  numbers.  The  temperature  at 
this  depth  is  below  zero  centigrade.  "Tout  un  monde 
de  bacteYies  existe  a  une  temperature  qui  descend  jns- 
qu'a  2°  an  dessous  de  ze>o." 

Coi).  GAZERT,  HANS.  Bakteriologische  Aufgaben 
der  deutschen  Sudpolar-Expedition.  Peter- 
martns  geogr.  Mitteil.,  Bd.  XLVII,  1901,  pp. 
I53-I55. 

Coi).  BELLI,  C.  M.  Chemische,  mikroskopische  und 
bakteriologische  Untersuchungen  iiber  den 
Hagel.  Hyg.  Rdsch.,  Berlin,  Bd.  xi,  1901, 
pp.  1,181-1,187. 

('02).  BINOT,  JEAN,  fitude  bacteriologique  du  massif 
du  mont  Blanc.  Nature,  Paris,  (ler  semest.), 
1902,  pp.  359-362,  av.  fig.  C.  R.  des  se.  de 
1'Acad.  des  sci.,  T.  cxxxiv,  1902,  pp.  673-676. 


256 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


XLI1I.    Soil-Organisms;  Putrefactive  Organisms. 

('82).  TYNDALL.    See  XLII. 

('86).  ADAMETZ.    See  XLII. 

('86).  BEUMER.    See  XLII. 

('87).  FRAENKEL.     See  XLII. 

('89).  REIMERS.    See  XLII. 

('91).  BERTHELOT,  M.,  ET  ANDRE,  G.  Sur  1'odeur 
prppre  de  la  terre.  C.  R.  de  se.  1'Acad.  des 
sci.,  Paris,  1891,  T.  cxn,  598. 

('91).  MANFREDI,  LUICI.  Sulla  contaminazione  della 
superficie  stradale  nelle  grandi  citta  dal 
punto  di  vista  dell'igiene  e  dell'ingegneria 
sanitaria.  Recerche  e  studi  fatti  con  speciale 
riguardo  alia  citta  di  Napoli.  Atti  della  R. 
Accad.  delle  sci.  fis.  e  mat.  di  Napoli.  2a 
serie,  vol.  IV,  1891,  appendice,  No.  4,  pp.  1-79. 

('93).  DUCLAUX.    See  XLII. 

('96).  DUCLAUX.    See  xxvin. 

('99).  LEVIN.    See  XLII. 

('01).  SMITH,  R.  GREIG.  Bacteria  and  the  disintegra- 
tion of  cement.  Proc.  Linn.  Soc.  of  New 
South  Wales,  vol.  xxvi,  for  the  year  1901, 
Part  i,  Sydney,  1902,  pp.  107-117.  Also  a 
separate  (issued  Aug.  13,  1901). 
Disintegration  not  due  to  the  bacteria. 

('02).  KATAYAMA,  T.  On  the  general  occurrence  of 
Bacillus  methylicus  in  the  soil.  Bull,  of  the 
College  of  Agr.,  Tokyo  Imperial  Univ.,  vol. 
v,  No.  2,  1002,  pp.  255-258.  Also  a  separate. 

('02).  CHESTER,  FREDERICK  D.  The  bacteriological 
analysis  of  soils.  Proc.  23d  Ann.  meeting 
of  Soc.  for  Prom.  Agric.  Sci.,  1902,  pp.  173- 
182.  Also  a  separate. 

('02).  CHESTER,  FREDERICK  D.  Bacteria  of  the  soil 
in  their  relation  to  agriculture.  Bulletin  No. 
98,  Dept.  of  Agric.  of  Pennsylvania,  1902, 
pp.  88,  with  plates.  A  bibliography  of  105 
titles. 

('02).  REMY,  TH.  Bodenbakteriologische  Studien. 
Centralb.  Bakt.,  Abt.  2,  Bd.  vm,  1902,  .pp. 
657-662,  pp.  699-705,  pp.  728-735,  pp.  761-769. 

('04).  CHESTER,  FREDERICK  D.  Observations  on  an 
Important  Group  of  Soil  Bacteria.  Organ- 
isms related  to  Bacillus  .subtilis.  Fifteenth 
Annual  Report  of  the  Delaware  College 
Agrl.  Exp.  Sta.,  for  1903,  Newark,  Del.,  U. 
S.  A.  With  5  plates.  Also  a  separate,  pp. 
1-54.  Copy  of  separate  received  from  author 
October  15,  1904. 


XLIV.    Vinegar-Bacteria. 

('61).  PASTEUR.  Acetic  fermentation  due  to  bac- 
teria. Ann.  scient.  de  1'Ecole  normale  supe- 
rieure,  1861. 

Not  seen. 

('68).  PASTEUR,  Louis.  Etudes  sur  le  vinaigre,  sa 
fabrication,  ses  maladies,  moyens  de  les 
prevenir;  nouvelles  observations  sur  la  con- 
servation des  vins  par  la  chaleur.  Paris, 
1868.  Gauthier-Villars,  Imprimeur-Libraire. 
Victor  Masson  et  Fils,  Libraires,  pp.  vm,  1 19. 

('86).  BROWN.    See  XLI. 


('86). 
('87). 
(93). 
('93). 


('94). 
('95). 

('97). 
('98). 

Coo). 


BROWN,  A.  J.  The  chemical  actions  of  pure 
cultivations  of  Bacterium  aceti.  Jour.  Chem. 
Soc.  Trans.,  1886,  vol.  XLIX,  London,  pp. 
172-187. 

BROWN,  A.  J.  Further  notes  on  the  chemical 
action  of  Bacterium  aceti.  Jour.  Chem.  Soc., 
London,  1887,  vol.  LI,  Transactions,  pp.  638- 
642. 

HANSEN,  EMIL  CHR.  Botanische  Unter- 
suchungen  iiber  Essigsaurebakterien.  Ber. 
d.  deutsch.  >bot.  Gesellsch.,  Bd.  xi,  1893,  pp. 
(69) -(73).  General  Versammlungs-Heft. 

LAFAR,  F.  Physiologist-he  studien  iiber  Essig- 
garung  und  Schnell-Essigfabrikation.  Cen- 
tralb. f.  Bakt.,  1893,  Bd.  xni,  pp.  684-697. 
Bibliography  of  13  titles. 

HANSEN,  E.  C.  Recherches  sur  les  bacteries 
acetifiantes.  Compt.  rend.  d.  trav.  du  Lab. 
Carlsberg,  T.  in,  Liv.  3,  pp.  182-216. 

HANSEN,  EMIL  CH.  Recherches  sur  les  bac- 
teries acetifiantes.  Ann.  de  micrographie, 
T.  vi,  1894,  No.  8,  pp.  385-395;  No.  9,  pp. 
441-470.  Also  a  separate,  pp.  41.  14  text  figs. 

LAFAR.  Physiologische  studien  iiber  Essig- 
garung  und  Schnellessigfabrikation.  Cen- 
tralb. f.  Bakt.,  2  Abt.,  Bd.  I,  1895,  pp.  129- 
ISO. 

LAFAR.     See  in. 

BEYERINCK,  M.  W.  Ueber  die  Arten  der  Es- 
sigbakterien.  Centralb.  f.  Bakt.,  2  Abt.,  Bd. 
iv,  1898,  pp.  209-216. 

HANSEN,  EMIL  CH.  Recherches  sur  les  bac- 
teries acetifiantes.  (Troisieme  memoire.) 
C.  R.  des  travaux  du  laboratoire  de  Carls- 
berg,  T.  v,  ire  Livraison,  1900,  pp.  39-46,  I 
fig.  Also  a  separate.  Copenhagen,  1900. 


XLV.     Silage-Bacteria,    Fermentation    of   Tobacco, 

of  Indigo,  Retting  of  Flax,  of  Sisal  Hemp,  Etc., 

Softening  of  Pickles,  Sauerkraut,  Etc. 

(See  also  XX  and  XLIV.) 

("87).  ALVAREZ,  E.  Sur  un  nouyeau  microbe,  determ- 
inant la  fermentation  indigotique  et  la  pro- 
duction de  1'indigo  bleu.  C.  R.  des  se.  de 
1'Acad.  des  sci.,  Paris,  1887,  T.  cv,  pp.  286- 
289. 

('89).  BURRILL,  T.  J.  The  biology  of  ensilage.  Bull. 
Ag.  Exp.  St.  Univ.  of  111.,  1889,  No.  vn, 
pp.  I77-I94. 

('91).  ALBERT,  FRIEDRICH.  Untersuchungen  iiber 
Grunpressf utter.  Jahrb.  d.  deutsch.  Land- 
wirtsch. -Gesellsch.,  Bd.  vi,  Tl.  i,  pp.  149- 
250,  Berlin,  1891. 

This  author  says  bacteria  exert   a  preponderant  influ- 
ence on  the  course  of  the  fermentation. 

('91).  SUCHSLAND,  EMIL.  Ueber  Tabaks  fermenta- 
tion. Ber.  d.  deutsch.  'hot.  Gesselsch.,  Bd. 
ix,  Berlin,  1891,  pp.  79-81. 

('94).  VAN  LOOKEREN-CAMPAGNE,  C.  J.  Bericht  iiber 
Indigo-Untersuchungen,  ausgefiihrt  an  der 
Versuchs-Station  zti  Klatten  auf  Java.  D. 
landw.  Vers.-Stat,  1894,  Bd.  XLIII,  pp.  401- 
426. 


SILAGE-BACTERIA,    FERMENTATION    OF   TOBACCO,    INDIGO,    FLAX,    ETC.          257 


('95)-  WINOGRADSKY,  SERGius.  Sur  le  rouissage  du 
lin  et  son  agent  tnicrobien.  C.  R.  des  se,  de 
1'Acad.  des  sci.,  Paris,  1895,  T.  cxxi,  pp. 
74^-745- 

A  r£sum£  of  the  principal  results  of  work  doue  by 
Fribes  iu  Wiuogradsky's  laboratory. 

1.  Stems  sterilized  under  water'by  a  short  heating  at 
100°,  repeated  three  days  in  succession,  or  by  one  heating 
at  115°  for  fifteen  minutes,  did  not  become  retted. 

2.  Sterilized   flax  immersed  in  water  and  inoculated 
with  any  one  of  the  maerobic  and  anaerobic  organisms, 
first  isolated  from  the  macerations  by  means  of  gelatin 
plates,  did  not  undergo,  even   after  several  mouths,  a 
commencement  of  retting,  nor  was  any  liberation  of  gas 
noticed.    On  the  contrary — 

3.  If  into  tubes  of   water  containing  sterilized  flax  a 
small  hit  of  straw  of  misterilized   flax  was  thrown    a 
very  active  fermentation  commenced  at  the  end  of  12  to 
15  hours,  and  at  the  end  of  two  or  three  days  the  retting 
was  completed. 

The  specific  organism  was  obtained  for  study  from  suc- 
cessive cultures  upon  steam -sterilized  flax,  protected 
from  the  air  by  immersion  in  deep  tubes  full  of  water, 
whose  surface  was  covered  by  a  layer  of  oil.  After  a  long 
enough  series  of  re-sowing  under  these  same  conditions, 
the  microscopic  study  of  these  cultures  has  removed  all 
doubt  about  the  agent  of  this  fermentation.  It  is  found 
almost  pure  in  the  interior  of  the  stem,  and  Fribes  has 
succeeded  iu  isolating  it  in  a  completely  pure  state  by 
cultivating  it,  in  the  absence  of  air,  upon  slices  of  cooked 
potato  rubbed  with  chalk.  It  is  a  bacillus  relatively 
large,  forming  spores  in  the  terminal  swellings  (tadpole 
form).  In  the  young  state  its  rods  are  from  10  to  15  m 
long,  with  a  th'ickuess  of  0.8  M;  often  one  finds  articu- 
lated filaments  much  longer  ;  thev  become  later  a  little 
thicker  { i  ju),  and  form  then  ovoid  swellings  3  ^  long  by 
2  n  thick  ;  the  ovoid  spore  which  is  formed  there  is  1.8  j* 
by  1.2  ft.  Sterilized  flax  was  retted  in  pure  cultures  of 
this  bacillus,  and,  after  undergoing  the  successive  opera- 
tions of  grinding  (dressing)  peeling  and  combing,  yielded 
a  fine  silky  flax  of  light  color  but  a  little  too  much  retted 
and  without  consistence. 

In  a  general  study  of  the  bacteria  of  retting,  Fribes  dis- 
covered the  following  facts  : 

1.  The  bacillus    ferments    glucose,  cane-sugar,  milk- 
sugar,  and  starch,  but  only  when  the  liquid  contains  pep- 
tone.   With  ammonia  as  the  only  source  of  nitrogen,  the 
bacillus  is  absolutely  void  of  action  on  these  eminently 
fermentable  substances. 

2.  Pectic  matters,  pectine,  or  pectic   acid,  extracted 
from    flax,  pears,  carrots,  white    turnips,  pure  as  they 
can  be  prepared,  are  decomposed,  in  presence  of  an  am- 
monium salt  as  the  sole  nitrogenous  food,  with  an  extra- 
ordinary facility. 

3.  Cellulose,  under  the  form  of  Swedish  filter  paper, 
or  as  an  amorphous  precipitate,  can  absolutely  not  he 
attacked  by  this  bacillus.     Gum  arable  is  not  fermented. 

4.  Vegetable    substance,  from  flax,  white  turnips,  ex- 
tracted cold  by  pure  water  and  water  slightly  acid  and 
alkaline,  and  submitted  to  fermentation  by  this  bacillus, 
loses  the  greater  part  of  the  substances  which  are  esti- 
mated as  pectic  matter  ;  also  the  loss  of  weight  of  the 
fermented  substance  corresponds  sensibly  to  the  content 
in  pectic  matter  of  the  nufermented  substance. 

As  a  result  of  these  studies  Winogradsky  concludes  that 
the  retting  of  flax  may  be  considered  as  a  pectic  fermen- 
tation in  the  micro-biological  sense  of  the  word,  of  which 
the  bacillus  described  is  the  specific  agent. 

1/96).  TOI,OMEI,  Giuiyio.  Ueber  die  Fermentation  der 
Oliven  und  die  Oxydation  des  Olivenoles. 
Atti  R.  Acad.  dei  Lincei  Roma,  se.  v,  Rendi- 
conti,  Classe  sci.  fis.,  matem.  e  nat.,  vol.  V, 
Feb.  16,  1896,  pp.  122-129. 
Due  to  an  enzyme.  Not  bacterial. 

('96).  VAN  LOOKEREN-CAMPAGNE,  C.  J.,  y.  VAN  DER 
VEEN,  P.  J.  Ueber  Indigpbildung  aus 
Pflanzen  der  Gattttng  "Indigo  fera."  D. 
landw.  Vers.-Stat.,  1896,  Bd.  XLVI,  pp.  249- 
258- 

('97).  CONRAD,  EUGEN.  Bakteriologische  und  chem- 
ische  studien  iiber  Sauerkrautgahrung. 
Arch.  f.  Hyg.,  Bd.  xxix,  1897,  pp.  56-9$- 
See  also  Zeitschr.  f.  Spiritusindustrie,  xx 
Jahrg.,  1897,  No.  23,  p.  188;  No.  24,  pp. 

2OO-2OI. 


The  fermentation  of  the  "  Weisskraut  "  is  attributed  to 
Bacterium  brassica;  acidse,  Lehm.  £  Conrad,  nearly  re- 
lated to  Bacillus  coli.  This  organism  is  motile,  grows 
aerobically  and  anaerobically,  produces  acids,  carbon 
dixoide,  hydrogen  and  marsh  gas.  Gelatin  is  not  liquefied. 
The  surface  colonies  are  gray  white  to  gray  yellow  on 
gelatin  and  agar,  and  are  bright  yellow  on  potato.  It 
ferments  maltose,  lactose  and  dextrose.  Most  of  the  acid 
is  lactic  acid. 

('98).  BREAUDAT,  L.     Sur  le  mode  de  formation  de 

I'indigo   dans   les   procedes   d'extraction  in- 

dustriels.        Fonctions      disastasiques      des 

plantes  indigpferes.    C.  R.  des  se.  de  1'Acad. 

des  sci.,  Paris,  1898,  T.  cxxvn,  pp.  769-771. 
('98).  MOLISCH.     Ueber  die  sogenannte  Indigogahr- 

ung  und  neue  Indigopflanzen.     Sitzungsber. 

d.  k.  Akad.  der  Wiss.  in  Wien,  July,  1898. 

Title  only. 

('98).  PRESCOTT  AND  UNDERWOOD.    See  xx. 
('99).  BREAUDAT.     Nouvelles  recherches  sur  les  fonc- 

tions  diastasiques  des  plantes  indigoferes.   C. 

R.    des    se.    de    1'Acad.    des    sci.,    Paris,   T. 

cxxvni,  1899,  pp.  1,478-1,480. 
('99).  VERNHOUT,  J.   H.     Onderzoek  over  bacterien 

bij  de  fermentatie  der  tabak.  Mededeelingen 

uit  s'Lands  Plantentuin,  xxxiv,  Batavia,  G. 

Kolff  &  Co.,  1809,  p.  49,  2  plates. 

The  fermentation  of  tobacco  is  ascribed  to  Bacillus 
tabaci-fermentationis  Vernhout. 

('99).  LoEw,  OSCAR.  Curing  and  fermentation  of 
cigar  leaf  tobacco.  U.  S.  Dept.  of  Agric., 
Report  No.  59,  Div.  Veg.  Phys.  &  Path., 
34  PP- 

('oo).  LOEW,   OSCAR.      Sind    Bakterien  die   Ursache 
der  Tabakfer.menta'tion  ?    Centralb.  f.  Bakt., 
2  Abt.,  Bd.  vi,  1900,  pp.  108-112. 
The  fermentation  is  strictly  enzymic,  and  bacteria  are 
not  concerned  in  it. 

('oo).  LoEW,  OSCAR.  Physiological  Studies  of  Con- 
necticut leaf  tobacco.  U.  S.  Dept.  of  Agric., 
Div.  of  Veg.  Phys.  &  Path.,  1900.  Report 
65.  57  PP- 

Coo).  BEIJERINCK.  Verdere  onderzoekingen  over  de 
indigovorming  uit  weedl  (Isatis  tinctoria). 
Proc.  K.  Akad.  Wetensch.  Amsterdam,  Deel 
ix,  June  30,  1900,  pp.  74-90.  Abstr.  in  Bot. 
Zekung,  2  Abt.,  vol.  58,  1900,  col.  188-189. 

The  production  of  indigo  blue  is  due  to  the  action  of 
the  enzyme  isatase  upon  isatan.  Isatan  occurs  m  the 
protoplasm  ;  isatase  occurs  in  the  chromatophores.  In 
the  living  cell  the  author  thinks  reactions  are  prevented 
by  the  acidity  of  the  cell-sap.  He  says  :  The  action  of 
isatase  upon  isatan  is  possible  only  in  neutral  or  ampho- 
teric  and  very  weakly  acid  solutions."  A  temperature  of 
48°  to  50°  C.  is  the  optimum  for  this  action. 

Coi)  LOEW,  OSCAR.  Catalase,  a  new  enzym  of  gen- 
eral occurrence,  with  special  reference  to  the 
tobacco  plant.  U.  S.  Dept.  of  Agric.  Rep. 
69,  Div.  Veg.  Phys.  &  Path.,  1901,  47  PP- 
('oi).  RUSSELL,  H.  L.,  AND  BABCOCK,  S.  M.  Concern- 
ing the  theories  of  silage  formation.  Science, 
n.  s.,  vol.  xni,  p.  328,  1901. 

The  conclusion  reached  is  that  bacteria  do  not  play 
any  very  considerable  role  in  the  fermentation  of  silage. 

('oi)    PREYER,     AXEL.      Ueber     Kakaofermemation. 

Tropenpflanzer,  Berlin,  Bd.  v,  1901,  pp.  157- 

173. 
('oi)    SCHULTE  IM  HoFE,  A.     Zur  Kakao-Fermenta- 

tion.     Tropenpflanzer,   Berlin,   1901,   Bd.   v, 

pp.  225-227. 


258 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('02).  BABCOCK,  S.  M.,  UND  RUSSELL,  H.  L.  Die  bei 
der  Herstellung  von  Garfutter  (Silage) 
wirkenden  Ursachen.  Centralb.  f.  Bakt.,  Abt. 
2,  Bd.  ix,  1902,  pp.  81-88. 

('02).  HAUMAN,  L.  fitude  microbiologique  et 
chimique  du  rouissage  aerobic  du  lin.  Ann. 
de  1'Inst.  Pasteur,  T.  xvi,  1902,  pp.  379-385. 

('02).  BAIL,  OSKAR.  Untersuchung  einiger  bei  der 
Verwesung  pflanzlicher  Stoffe  thatiger 
Sprosspilze.  Centralb.  f.  Bakt.,  Abt.  2,  Bd. 
vin,  1902,  pp.  567-584. 

('03).  WEHMER,  C.  Die  Sauerkrautgiirung.  Cen- 
tralb. f.  Bakt.,  2  Abt.,  1903,  x  Bd.,  pp.  625-629. 

('04).  STOERMER,  K.  Ueber  die  Wasserroste  des 
Flachses.  Ceiitralblatt  f.  Bakt.,  2  Abt.,  Bd. 
XHI,  No.  1-3,  Sept.,  1904,  pp.  35-45-  Con- 
tinued. 

('04).  BEIJERINCK,  M.  W.,  AND  VAN  DELDEN,  A.  "On 
the  bacteria  which  are  active  in  flax-ret- 
ting." Koninklijke  Akademie  van  Weten- 
schappen  te  Amsterdam.  Proceedings  of 
the  Meeting  of  Jan.  30,  1904.  Pub.  Feb.  25, 
1904,  pp.  462-481,  i  .plate.  Also  a  separate. 

('04).  OM£LIANSKI,  W.  Die  histologischen  und 
chemischen  Veranderungen  der  Leinstengel 
unter  Einwirkurg  der  Mikroben  der  Pektin- 
und  Cellulosegarung.  Centralb.  f.  Bakt.,  2 
Abt.,  xii  Bd.,  igo4,  pp.  33-43,  i  plate. 

('04).  WEHMER,  C.  Die  Sauerkrautgaerung.  Be- 
richt  des  V.  Internationale!!  Kongresses  f. 
angewandte  Chemie  zti  Berlin,  1903.  Sekt. 
VI,  Bd.  in,  Berlin,  1904,  p.  712.  Also  a 
separate,  5  pp. 

Obtains  results  decidedly  different  from  Conrad.  The 
ordinary  sauerkraut  fermentation  is  a  mixed  fermenta- 
tion. Gas-development  is  due  to  a  bottom  yeast;  acid 
production  to  a  non-liquefying,  non-motile,  non-gas- 
fonniug,  lactic  acid  bacterium. 


XLVI.    Bacteria  in  Water  and  Ice;  Dung-Bacteria. 

(See  also  XXXIII  and  XXXIV.) 

('76).  WARMING.     See  xn. 

('86).  FRAENKEL,  CARL.  Ueber  den  Bakteriengehalt 
des  Eises.  Zeitschr.  f.  Hyg.,  Bd.  i,  1886, 
pp.  302-314. 

(87).  MACE.  >Sur  quelques  bacteries  des  eaux  de 
boisson.  Ann.  d'hyg.  publ.  et  de  med.  leg., 
avril,  1887.  3d  series,  T.  xvn,  pp.  354-357- 
(87).  PRUDDEN,  T.  MITCHELL.  On  Bacteria  in  ice 
and  their  relation  to  disease,  with  special 
reference  to  the  ice  supply  of  New  York 
City.  Med.  Record,  March  26  and  April  8, 
1887,  Nos.  13  and  14;  also  a  separate,  61  pp. 
Reviewed  in  Centralb.  f.  Bakt.,  Bd.  I,  1887, 
pp.  650-652,  and  Ann.  de  J'Inst.  Pasteur,  T.  I, 
1887,  pp.  409-410. 

Dr.  Prudden  tested  tlie  resistance  of  various  bacteria  to 
prolonged  cold,  in  blocks  of  ice  and  to  repeated  freezings 
and  thawiugs.  Proteus  vulgaris  and  Bacillus  prodigiosus 
did  not  grow  after  51  days  freezing.  A  slender  liquefying 
bacillus  from  Crotou  water  was  killed  in  seven  days 
The  following  withstood  freezing  :  Staphylococcus  pyo- 
genes  (66  days)  ;  a  fluorescent  bacillus  from  ice  (77 
days)  ;  bacillus  of  typhoid  fever  (123  days)  cultures  made 
at  intervals  showed  less  and  less  living,  but  all  were  not 
destroyed.  Repeated  freezings  and  thawings  were  more 
fatal  to  the  typhoid  bacillus  than  a  constant  low  tem- 
perature. Five  freezings  and  thawings  at  intervals  of 
three  days  destroyed  tliis  bacillus. 

('87).  BORDONI-UFFREDUZZI,  GuiDO.  Die  biologiscbe 
Untersuchung  des  Eises  in  seiner  Beziehung 
zur  offentlichen  Gesundheitspflege.  Cen- 
tralb. f.  Bakt.,  1887,  Bd.  H,  pp.  489-497. 


('88).  SCHMELCK,  L.  Eine  Gletscherbakterie.  Cen- 
tralb. f.  Bakt.,  1888,  Bd.  iv.  pp.  545-547. 

A  green  fluorescent  organism  was  the  commonest  form 
This  was  a  short  rod  which  liquefied  gelatin. 

('91).  VIRON,  L.  Du  role  des  Schizophytes  dans  les 
reactions  qui  se  passent  dans  les  eaux  dis- 
tillees.  Jour,  de  Pharm.  et  de  Chini.,  1891, 
T.  xxm,  series  5,  pp.  586-593. 

('91).  NORDT MEYER.    See  xvn. 

('94).  LASER.     See  xx. 

('94).  HOUSTON.    See  xvn. 

('94).  FRANKLAND,  P.,  AND  MRS.  P.  Micro-organ- 
isms in  water.  London,  1884,  pp.  xr,  532. 

('95).  SEDGWICK  UND  PRESCOTT.  On  the  influence  of 
variations  in  the  composition  of  nutrient 
gelatin  upon  the  development  of  water  bac- 
teria. Am.  Pub.  Health  Asso.,  vol.  xx,  1895, 
pp.  450-458.  Rev.  in  Centralb.  f.  Bakt.,  xix 
Bd.,  1896,  p.  222. 

('95).  RABINOWITSCH.     See  xxxiv. 

('95)-  SMITH,  THEOBALD.  Notes  on  bacillus  coli 
communis  and  related  forms;  together  with 
some  suggestions  concerning  the  bacterio- 
logical examination  of  drinking  water.  The 
Amer.  Journal  of  the  Med.  Sci.,  Sept.,  1895. 
Also  a  separate,  pp.  20. 

('95).  SMITH,  THEOBALD.  Ueber  den  Nachweis  des 
Bacillus  coli  communis  im  Wasser.  Cen- 
tralb. f.  Bakt.,  xvin  Bd.,  1895,  pp.  494-495. 

('95).  SEVERIN,  S.  A.  Die  im  Miste  vorkommenden 
Bakterien  und  deren  physiologische  Rolle 
bei  der  Zersetzung  derselben.  Centralb.  f. 
Bakt.,  2  Abt.,  Bd.  I,  1895,  pp.  97-114  and 
709-817. 

('97).  HESSE,  FRIEDR.  Ueber  die  Verwendung  von 
Nahragar-Agar  zu  Wasseruntersuchungen. 
Centralb.  f.  Bakt.,  xxi  Bd.,  1897,  pp.  932-937. 

Finds  agar-media  better  than  gelatin  on  account  of 
number  of  liquefying  organisms  constantly  present  in 
water. 

('97).  KERN,  HEINRICH.  Beitrag  zur  Kenntniss  der 
im  Darme  und  Magen  der  Vogel  vorkom- 
menden Bacterien.  Arb.  a.  d.  bact.  I  list.  d. 
tech.  Hochschule  zu  Karlsruhe,  Bd.  i,  Heft 
iv,  1897,  pp.  379-532. 

Many  bacteria  are  described  at  length. 

CgS).  WARD,    H.    MARSHALL.      Some    Thames    Bac- 
teria.    Annals  of  Botany,  vol.  xn,  1898,  pp. 
287-322.     Two  double  plates  in  color. 
This  paper  treats  of  (i)  A  short  colorless  bacterium 
forming  stearine-like  colonies  :  type  of  Bacterium  ureae 
(Jaksch);  (2)  A  colorless  capsuled  coccus  or  bacterium  ; 

(3)  rose-pink  Micrococcus  :  type  of  M.  carneus  (Zimni.)  ; 

(4)  A  pseudo-bacillus. 

('99).  KASANSKY.    See  xxxm. 

Cc9).  FULLER,  GEORGE  W.,  AND  JOHNSON,  GEORGE  A. 
On  the  differentiation  and  classification  of 
water  bacteria.  Jour.  Exp.  Med.,  vol.  iv, 

1899,  PP-  609-626;  also  a  separate. 

Coo).  KOHLBRUGGE,  J.  H.  F.  Vibrion-Studieii.  i. 
Die  Ubiquitat  choleraahnlicher  Wasser- 
vibrionen.  Centralb.  f.  Bakt.,  xxvm  Bd., 

1900,  pp.  721-726.     it.   Panmorphismus  und 
erbliche  Variationen.     Ibid,  pp.  833-842. 

Coo).  SEUGWICK  AND  WINSLOW.    See  xxxm. 

Coi).  PARK  .See  xxxm. 

Coi).  HORHOCKS,  W.  H.  An  introduction  to  the  bac- 
teriological examination  of  water.  London, 
J.  &  A.  Churchill,  1901,  pp.  x,  300,  5  plates. 


MILK-HACTERIA,    BUTTER-BACTERIA,    CHEESE-BACTERIA,    MEAT-BACTERIA.      259 


('01).  GAGE,  STEPHEN  DE  M.  Bacteriological  Studies 
at  the  Lawrence  Experiment  Station,  with 
special  reference  to  the  determination  of  B. 
coli.  33d  An.  Rep.  St.  Bd.  of  Health  of 
Mass.,  for  1901,  pp.  397-420.  Also  a  sepa- 
rate, pp.  26. 

Coi).  Micur.A,  W.  Compendium  der  bakteriologis- 
chen  Wasseruntersuchung  nebst  vollstaen- 
diger  Ucbersicht  dcr  Trinkwasserbakterien. 
O.  Nemnich,  Wiesbaden,  1901,  pp.  vn,  440, 
with  2  plates. 


('02). 

('03). 


Coj). 


(03). 

(03). 
('04). 
(•04). 


MACFADYEN.    See  xxxm. 

JORDAN,  EDWIN  OAKES.    The  kinds  of  Bacteria 

found  in  river  water.     Journal  of  Hygiene, 

vol.   in,  No.   i,   1903.     Also  a  separate,  pp. 

1-27. 
IMMENDORF,    H.     Ueber    Stallmist-Bewahrung 

(Konservierung)    mil    Chemischen    Mitteln. 

Berlin,  Mitt.  d.  Landw.  Ges.,  Bd.  xvm,  1903. 

pp.  99-101. 
SCHUEDER.     See  xvm. 

WlNSLOW   AND   NlBECKER.      See   XVII. 

GAGE  AND  ADAMS.    See  xvi. 
STOKES.    See  xvm. 


XLVII.     Milk-Bacteria;  Butter-Bacteria;  Cheese- 
Bacteria;  Meat-Bacteria. 

C8i).  JALAN  DE  LA  CROIX.    See  xxxvm. 

I'Sj).  ScHMiDT-MuEHLHEiM.  Untersuchungeti  iiber 
fadenziehende  Milch.  Pfliiger's  Archiv., 
1882,  Bd.  xxvii,  pp.  490-510,  i  fig. 

('84).  HuEi'PE,  FERDINAND.  Untersuchungen  iiber 
die  Zersetzungen  der  Milch  durch  Micro- 
organismen.  Mitth.  a.  d.  K.  Gesundheits- 
amte,  Bd.  II,  Berlin,  1884,  pp.  309-371. 

('89).  MENGE,  KARL.  Ueber  rothe  Milch.  Centralb. 
f.  Bakt.,  vi  Bd.,  1889.  pp.  596-602. 

('89).  BAGINSKY,  ADOLF.  Rote  Milch.  Deutsche 
Medizinal-Zeitung  ,1889,  No.  9,  pp.  106-107. 

('89).  BAGINSKY,     ADOLF.       Zum     Grotenfelt'schen 

Bacillus  der  roten  Milch.     Deutsche  mediz. 

Wochenschrift,  1889,  Bd.  xv,  p.  212. 

Tliis   organism  was  isolated   from    feces.     It  liquefied 

gelatin  slowly  and  colored  milk  a  dirty  red  or  red-brown. 

('91).  CONN,  H.  W.  Ueber  einen  bittere  Milch 
erzeugenden  Micrococcus.  Centralb.  f.  Bakt., 
ix  Bd.,  1891,  pp.  653-655. 

('91).  ADAMETZ,  L.  Untersuchungen  iiber  Bacillus 
lactis  viscosus,  einen  weitverbreketen  milch- 
wirthschaftliohen  Schadling.  Berliner  land- 
wirthschaftlichc  Jahrbiicher,  1891,  Bd.  XX, 
pp.  185-207,  i  plate.  Rev.  in  Centralb.  f. 
Bakt.,  ix  Bd.,  1891,  pp.  698-700. 

('9-0.  NENCKI,  L.,  ET  ZAWADZKI,  J.     Sur  la  sterilisa- 
tion du  lait.     Arch,   des   Sci.   Biol.  publiees 
par  L'institut  Imperial   de  Med.  Exp.  a  St. 
Petersbourg,  T.  I,  1892,  pp.  371-397. 
Contains  a  bibliography  of  50  numbers. 

('93).  DUCLAUX,  E.  Sur  le  role  protecteur  des 
microbes  dans  la  creme  et  les  fromages. 
Ann.  de  I'lnst.  Pasteur,  T.  vn,  1893,  pp. 
305-324- 

('93).  BLEISCH,  MAX.  Ueber  bittere  Milch  mid  die 
Sterilisierung  der  Milch  durch  Erhitzen 
unter  Luftabschluss.  Zeitschr.  f.  Hyg.,  1893, 
Bd.  xin,  pp.  81-99. 


('94).  BORDONI-UFFREDUZZI,    GUIDO.      Ein    Fall    von 

fuchsinahnlicher        Bakterienfarbung        des 

Fleisches.      Hygien.    Rundschau.,    1894,    Bd. 

iv,  pp.  12-14. 
('94).  v.  KLECKI,  VALERIAN.     Ueber  einige  aus  ran- 

ziger    Butter    kultivierte    Mikroorganismen. 

Centralb.  f.  Bakt.,  Bd.  xv,  1894,  pp.  354-362. 
('94).  LEICHMANN,  G.   Ueber  eine  schleimige  Garung 

der    Milch.      Landw.    Ver.-Stat.,    Bd.    XLIII, 

1894,  pp.  375-398.    Rev.  in  Centralb.  f.  Bakt., 

Bd.  xvi,  1894,  pp.  122-123. 
('94).  RussELL,  H.  L.   Outlines  of  dairy  Bacteriology, 

pp.  vi,  186,  1894.     Pub.  by  author.     Madison, 

Wisconsin. 
('94).  PAMMEL,  L.  H.    An  aromatic  bacillus  of  cheese 

(Bacillus  aromaticus  n.  sp.).    Extracts  from 

the   Iowa   Agricultural    Exp.    Station,    Bull. 

No.  21,  1894,  pp.   1-5.     Rev.  in  Centralb.  f. 

Bakt.,  Bd.  xvi,  1894,  p.  128. 
('94).  WEIGMANN,  H.,  UND  ZIRN,  GG.   Ueber  "seifige" 

Milch.     Centralb.  f.  Bakt.,  Bd.  xv,  1894,  pp. 

403-47O>  -mit  2  Abbildungen. 
('94).  DUCLAUX,  EMILE.     Le  lait;  etudes  chimiques 

et  microbiologit|iies.  2  tirage,  augmente   de 

notes   sur  le   role   des   microbes   et   sur  les 

phosphates   du  lait.     376  pp.,   I2mo.     Paris, 

J.  B.  Bailliere  &  fils.,  1894. 
('94).  HENRICI,  H.     Beitrag  zur  Bakterienflora   des 

Kiises.     Arb.  a.  d.  bact.  Inst.  d.  tech.  Hoch- 

schule   zu   Karlsruhe,   Bd.    i,   Heft    i,   1804, 

pp.  i-iio. 

('95).  RABINOWITSCH.     See  xxxiv. 
('95).  JOLLES,  MAX,  u.  WINKI.ER.  FERDINAND.     Bak- 

teriologische    Studien    iiber    Margarin    und 

Margarinproductes.     Zeitschr.    f.    Hyg.,   Bd. 

xx,  1895,  pp.  60-108. 

The  bacterial  content  of  margariu  products  is  slight  in 
comparison  with  that  of  natural  butter. 

('95).  CONN,  H.  W.  Bacteria  in  the  dairy,  vi.  Ex- 
periments in  ripening  cream  with  Bacillus 
-  No.  41.  7th  Ann.  Rep.  of  the  Storrs  Ag. 
Ex.  Station  for  1894,  pp.  57-68.  Middletown, 
Conn.,  1895. 

('96).  CONN,  H.  W.  The  relation  of  pure  cultures  to 
the  acid,  flavor,  and  arorna  of  butter.  Cen- 
tralb. f.  Bakt.,  2  Abt.,  Bd.  11,  1896,  pp.  409- 
415. 

('97).  CONN,  H.  W.  Butter  arorna.  Centralb.  f. 
Bakt.,  2  Abt.,  Bd.  ill,  1897,  pp.  177-179. 

('59).  MAASZEN,  ALBERT.  Fruohtatherbildende  Bak- 
terien.  Arb.  a.  d.  k.  Gesundheitsamte,  Bd. 
xv,  Berlin,  1899,  pp.  500-513,  3  pi.  from 
photomicrographs. 

The  plates  and  text  deal  largely  with  Bacillus  esterifi- 
caus,  B.  ester  fluoresceus  aim  B.  prjepolleus.  There  are 
numerous  references  to  literature. 

('59).  MOORE,  V.  A.,  AND  WARD,  A.  R.  An  inquiry 
concerning  the  source  of  gas  and  taint-pro- 
ducing bacteria  in  cheese  curd.  Bull.  No. 
158,  Cornell  Univ.  Agr.  Exp.  Station,  1899, 
pp.  221-237,  I  plate. 

('99).  WARD,  ARCHIBALD  R.     Ropiness  in  milk  and 
cream.     Cornell  Univ.  Agr.  Exp.  Sta.  Dairy 
Div.  Bui.  165,  1899,  pp.  395-412,  4  figs. 
Ropiness  attributed  to  Bacillus  lactisviscosus. 

('99).  STADLER.    See  xxxvui. 

Coo).  WEBER,    A.      Die    Bakterien    der    sogcnannten 

sterilisirten   Milch   des   Handles   ihre  biolo- 

gischen   Eigenschaften,   etc.     Arb.    a.   d.   k. 

Gesundheitsamte,  Bd.  xvn,  1900,  pp.  108-155. 

With  bibliography  of  225  titles. 


260 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('oi).  CHODAT,  R.,  ET  HOFMAN-BANG,  N.  O.  Les 
bacteries  lactiques  et  !eur  importances  dans 
la  maturation  du  fromage.  Ann.  de  1'Inst. 
Pasteur,  T.  xv,  1901,  pp.  36-48. 

Discusses  relation  of  tyrothrix  to  ripening  of  cheese. 

('oi).  WARD,  ARCHIBALD  R.  Further  observations 
upon  ropiness  in  milk  and  cream.  Cornell 
Univ.  Agr.  Exp.  Sta.  Dairy  Div.  Bull.  195, 
1901,  pp.  29-39,  2  figs. 

('oi).  PARK,  WM.  HALI.OCK.  The  great  bacterial 
contamination  of  the  milk  of  cities.  Can  it 
be  lessened  by  the  action  of  Health  authori- 
ties? 'The  Journ.  of  Hygiene,  vol.  I,  1901, 
pp.  391-406.  See  also  N.  Y.  Univ.  Bull,  of 
the  Med.  Sci.,  vol.  i,  1901,  pp.  7i-£6. 

('02).  FISCHER,  BERNHARD.  Zur  Aetiologie  der 
sogenannten  Fleisohvergiftungen.  Zeitschr. 
f.  Hyg.,  Bd.  xxxix,  1902,  pp.  447-510,  2 
plates.  With  a  bibliography  of  31  titles. 

('02).  EPSTEIN,  ST.  Untersuchungen  iiber  die  Reif- 
ung  von  Weichkasen.  Arch.  f.  Hyg.,  Bd. 
XLIII,  1902,  pp.  1-20. 

('02).  CONN,  H.  W.,  AND  ESTEN,  W.  M.    The  com- 
parative growth  of  different  species  of  bac- 
teria in  normal  milk.    Fourteenth  Ann.  Rep. 
Storrs    Agr.    Exp.    Station,    Storrs,    Conn., 
1001,  pp.  13-80.     Middletown,  Conn.,  1902. 
Relates  to  those  bacteria  occurring  naturally  in  milk 
and  not  to  those  introduced  by  the  bacteriologist. 

('02).  GRUBER,  TH.  Ueber  einen  die  Milch  rosafar- 
benden  Bacillus.  Bacillus  lactorubefaciens. 
Centralb.  f.  Bakt,  Abt.  2,  Bd.  vin,  1902,  pp. 
457-462. 

('02).  CRUDER,  TH.  Ueber  eine  in  der  Milch  Riiben- 
geruch  tind  Rubengeschmack  erzeugende 
Bakterie.  Molk.  Zeitung,  Hildesheim,  Bd. 
xvi,  1002,  pp.  351-353- 

('02).  HARDING,  H.  A.,  UNO  ROGERS,  L.  A.     Rost- 
flecken  in  Cheddarkase.     Centralb.  f.  Bakt., 
Abt.  2,  Bd.  vin,  1902,  pp.  442-443. 
Ascribed  to  Bacillus  rudensis. 

('02).  RICHTER,  ALBRECHT  P.  F.  Bakterielles  Ver- 
halten  der  Milch  bei  Boraxzusajtz.  Arch.  f. 
Hyg.,  Bd.  XLIII,  1902,  pp.  151-156. 

('02).  GRIMM,  MAX.  Ueber  einen  neuen  aroma- 
bildenden  Bacillus  nebst  einigen  Bemer- 
kungen  i\l>er  Reinkulturen  fur  Exportbutter. 
Centralb.  f.  Bakt.,  Abt.  2,  Bd.  VIH,  1902,  pp. 
584-590. 

('02).  ROSAM,  A.  Ueber  Konservierung  der  Milch 
mittels  WassersiofTsuperoxyd.  Centralb.  f. 
Bakt.,  Abt.  2,  Bd.  vin,  1902,  pp.  739-744,  pp. 
769-774. 

('03).  HARRISON,  F.  C.,  AND  GUMMING,  M.  The  bac- 
terial flora  of  freshly-drawn  milk,  Part  rv. 
Journal  of  Applied  Microscopy,  vol.  VI,  1903, 
No.  2,  p.  3,181.  Bibliography  of  25  titles. 

('03).    SWITHINBANK,  HAROLD,  AND  NEWMAN,  GEORGE. 

Bacteriology  of  Milk.  With  special  chapters 
•by  Dr.  Newman  on  the  spread  of  disease  by 
milk  and  the  control  of  the  milk  supply. 
With  chromo-lithographs,  Woodfoury  type 
reproductions  of  photographs  of  cultures, 
and  other  illustrations  of  bacteria  and  of 
apparatus,  and  also  charts  illustrating  epi- 
demics. London,  1903,  John  Murray,  pp. 
xx,  605. 


('03).  CONN,  H.  W.,  AND  STOCKING,  W.  A.,  JR.  Com- 
parison of  bacteria  in  strained  and  un- 
strained samples  of  milk.  Rep't  of  Storrs 
Agr.  Exp.  Station,  Conn.,  1902-3,  pp.  33-37. 

('03).  CONN,  H.  W.,  AND  STOCKING,  W.  A.,  JR. 
Series  n.  Strained  and  unstrained  milk 
preserved  at  70°  and  50°.  Rep't  of  Storrs 
Agr.  Exp.  Sta.,  Conn.,  1902-3,  pp.  38-51. 

('03).  CONN,  H.  W.,  AND  STOCKING,  W.  A.,  JR. 
Series  HI.  Aseptic  milk.  Rep't  of  Storrs 
Agr.  Exp.  Station,  Conn.,  1902-3,  pp.  52-62. 

('03).  CONN,  H.  W.,  AND  ESTEN,  W.  M.  Qualitative 
analysis  of  bacteria  in  market  milk.  Rep't 
of  Storrs  Agr.  Exp.  Station,  Conn.,  1902-3, 
pp.  63-91. 

('03).  CONN,  H.  W.  Bacteria  in  freshly  drawn  milk. 
Rep't  of  Storrs  Agr.  Exp.  Station,  Conn., 
1902-3,  pp.  92-98. 

('03).  CONN,  H.  W.  "The  relation  of  temperature  to 
the  keeping  property  of  milk."  Storrs  Agr. 
Exp.  Station,  Storrs,  Conn.  Bull.  26,  Oct., 

1903,  PP.  3-I5- 
The  author's  summary  is  as  follows  : 

1.  Variations  in  temperature  have  a  surprising  influ- 
ence upon  the  rate  of  multiplication  of  bacteria.    At  50° 
these  organisms  may  multiply  only  5-fold  in  24  hours, 
while  at  70°  they  may  multiply  7,so-fold. 

2.  Temperature  has  a  great  influence  upon  the  keeping 
property  of  milk.    Milk  kept  at  95°  (heat  of  the  cow's 
body)  will  curdle  in  18  honrs,  while  the  same  milk  kept 
at  70°  will  not  curdle  for  48  hours,  and  if  kept  at  50°  F. 
the  temperature  of  an  ice-chest,  may  sometimes  keep 
without  curdling  for  two  weeks  or  more. 

3.  So  far  as  the  keeping  property  of  milk  is  concerned, 
the  matter  of  temperature  is  of  more  significance  than 
the  original  contamination  of  the  milk  with  bacteria. 

4.  Milk  preserved  at  50°  or  lower  will  keep  sweet  for  a 
long  time,  but  it  becomes  filled  with  bacteria  of  a  more 
unwholesome  type  than  those  that  grow  at  higher  tem- 
peratures.   Old  milk  is  not  fit  for  market,  even  though  it 
be  perfectly  sweet. 

('03).  WILHELMY.     Die   Bakterienflora   der   Fleisch- 
extracte  und  einiger  verwandter  Praparate. 
Arb.  a.  d.  Bact.  Inst.  der  techn.  Hochschule 
zu  Karlsruhe,  in  Bd.,  i  Heft,  1903,  pp.  1-42, 
with  3  plates  (18  photomicrographs). 
Most  of  the  bacteria  exist  in  form  of  spores.    Twelve 
new  species  are  described:  Micrococcus  carniphilus,  M. 
margiuatus,  Streptococcus  debilis.  Bacterium  naveum,  B. 
insulosum,  Bacillus  carniphilus,  B.  canaliculatus,  B.  car- 
nis,  B.  iutertnittens,  B.  anthraciformis,  B.  glaciformis, 
B.  micans,  B.  kaleidoscopicus.   B.  carniphilus  occurs  in 
most  meat  extracts.     Other  species  found   were  :  Bacte- 
rium rusticum   Kern,  Bacillus  meseutericus  Fliigge,  B. 
vulgatus  Fliigge,  B.  cereus  Frankland,  B.   laevis  Frank- 
land,  and    B.  vegetus  Keni. 


XLVIII.  Bacteria  in  Bread. 

('85).  LAURENT,  EMILE.  La  bacterie  de  la  fermenta- 
tion panaire.  Bull,  de  1'Acad.  roy.  de  Bel- 
gique,  3  ser.,  T.  x,  1885,  pp.  765-775. 

(,'&&).  ARCANGEH,  G.  Sulla  fermentazione  panaria. 
Atti  della  Societa  toscana  di  scienze  natural! 
residente  in  Pisa.  Memorie.  Pisa,  1888,  vol. 
ix,  pp.  190-211.  Bibliog.  29  titles. 

('89).  KRATSCHMER.  UND  NIEMILOWICZ.  Ueber  eine 
eigentiirhliiche  Brotkrankheit.  Wiener  klin- 
ische  Wochenschrift,  1889,  Bd.  H,  pp.  593- 

594- 

Authors  ascribe  the  stringy  bread  to  Bacillus  mesen- 
tericus  vulgatus  Fliigge.  They  did  not  determine  from 
what  source  the  bread  was  infected,  but  state  that  the  or- 
ganism will  not  grow  in  acid  dough  or  acid  bread,  but 
that  it  grows  luxuriantly  in  feebly  alkaline  dough  or 
bread. 


BACTERIA  IN  BREAD;  IRON  BACTERIA;  SULPHUR  BACTERIA. 


26l 


('89).  PETERS,  W.  L.  Die  Organismen  des  Sauer- 
teiges  und  ihre  Bedeutung  fur  die  Brot- 
gahrung.  Bot.  Zeitung,  1889,  Bd.  XLVII,  col. 
405-419,  421-431,  437-449- 

('90).  UFFELMANN,  J.  Verdorbenes  Brot.  Centralb. 
f.  Bakt..  vin  Bd.,  1890,  pp.  481-485. 

(.'90).  POPOFF,  M.  Sur  un  bacille  anaerobie  de  la 
fermentation  panaire.  Ann.  de  1'Inst.  Pas- 
teur, 1890,  T.  iv,  pp.  674-676. 

("94).  LEHMANN,  K.  B.  Ueber  die  Sauerteiggarung 
und  die  Beziehungen  des  Bacillus  levans 
zum  Bacillus  coli  communis.  Centralb.  f. 
Bakt.,  Bd.  xv,  1894,  pp.  35O-354- 

('94).  WALDO,  F.  J.,  AND  WALSH,  DAVID.  Does  bak- 
ing sterilize  bread?  Tihe  Lancet,  London, 
1894  (n),pp.  906-908. 

The  general  conclusion  is  that  baking  does  not  fully 
sterilize.  The  authors  cultivated  numerous  kinds  of 
bacteria  from  the  interior  of  baked  loaves. 

('97)-  VOCEL,  J.     Beitrag  zur  Kenntniss  des  "faden- 

ziehenden   Brotes."     Zeitschr.    f.   Hyg.,   Bd. 

xxvi,  1897,  pp.  398-416. 
(*99)-  JUCKENACK,    ADOLF.      Beitrag    zur    Kenntniss 

des   "fadenziehenden   Brotes."     Zeitschr.    f. 

Untersuoh.    d.    Nahrungs-    und    Genussmit- 

teln,  ii  Jahrg.,  1899,  pp.  786-788. 
('oo).  THOMANN,    J.      Beitrag    zur    Kenntniss    des 

"fadenziehenden  Brotes."    Centralb.  f.  Bakt., 

2  Abt.,  Bd.  vi,  1900,  pp.  740-743. 

Stringy  bread  was  found  due  to  bacteria  introduced 
with  the  flour.  The  bacillus  isolated  by  the  author  out 
of  bread  and  two  kinds  of  flour  is  called  Bacillus  panis 
viscosi  (Vogel).  Its  cultural  characters  are  given  as 
follows  : 

It  is  sporiferous,  actively  motile,  stains  by  Gram's 
method,  liquefies  gelatin  rapidly,  forms  a  dry.  gray-white 
growth  on  agar,  spreads  widely  and  is  wrinkled  and  gray- 
white  on  potato,  grows  in  grape-sugar  bouillon  without 
gas-formation,  produces  a  thick  pellicle  on  peptone 
bouillon  with  a  clear  fluid  under  it,  and  grows  best  at 
40°  to  42°  C. 

('01).  DUCLAUX.     Pain  filant.     See  Traite,  T.  iv,  pp. 

513-SiS. 
('01).  BEULSHAUSEN,  FRIEDRICH.     Zur  Kenntnis  der 

Ursaohe     des     Klebrigwerdens     von     Brot. 

Diss.  Rostock  (Druck  v.  C.  Hinstorff),  1901, 

p.  24. 
('02).  MICHELS,    WOLFGANG.      Zur    Entsitehung    des 

fadenziehenden  Brotes.    Diss.  Kiel.  Konigs- 

berg  i.  Pr.     (Durck  v.  Hantung),  1902,  p.  15. 
('02).  LEHMANN,  K.  B.     Hygienische  Untersuchun- 

gen  iiber  Mehl  und  Brot.    x.   Neue  Studien 

iiber  die  Aciditat  des  Brotes,  ihre  Ursachen 

und  i'hre  beste  Bestimmungs-methode.     Arch. 

Hyg.,  Mtinchen,  Bd.  XLIV,  1902,  pp.  214-237. 


XLIX.    Iron-Bacteria. 

('88).  WINOGRADSKY,  S.  Ueber  Eisenbacterien.  Bot. 
Zeitung,  46  Jahrg.,  1888,  col.  261-270. 

('92).  MOLISCH,  HANS.  Die  Pflanze  in  ihrer  Bezie- 
hung  zum  Eisen.  Eine  physiologische  Studie. 
Jena,  1892,  Gustav  Fischer,  119  pp.,  i  table. 

('95).  MACALLUM,  A.  B.  On  the  distribution  of  as- 
similated iron  compounds,  other  than  -haemo- 
globin and  haematins,  in  animal  and  vege- 
table cells.  Quarterly  journal  of  microsc. 
science,  1895-06,  vol.  xxxvm,  new  series, 
No.  150,  pp.  175-274,  with  3  plates. 

The  part  relating  to  the  bacteria  begins  on  page  254. 


('97)-  MARPMANN,  G.  Bakteriolqgische  Mitteilungen. 
i.  Ueber  einen  neuen  Nahrboden  fur  Bak- 
terien.  n.  Ueber  ferrophile  Bakterien.  in. 
Ueber  den  Zusanimenhang  von  pathogenen 
Bakterien  mit  Fliegen.  Centralb.  f.  Bakt., 
xxn  Bd.,  1897,  pp.  122-132. 

The  new  substratum  is  raw  silk.  Author  has  found  a 
bacterium  which  stores  iron  in  its  cell-contents.  It  is 
non-motile,  2-3  x  0.8-2.0 >»,  ends  rounded,  form  plump,  with 
black  polar  chromatophores  and  intermediate  gray  gran- 
ules. Many  cells  are  entirely  black  and  opaque.  The 
pigment  is  insoluble  in  alcohol,  ether,  carbon  bisulphid 
and  benzine.  It  becomes  bluish  opalescent  with  am- 
monia, and  bleaches  with  HC1,  giving  off  hydrogen  sul- 
phid.  On  adding  ferricyauide  of  potash  after  HC1,  or 
with  it,  the  bacteria  become  an  intense  blue.  On  pep- 
tone-gelatin the  organism  was  white,  but  when  a  trace 
of  iron  sulphate  was  added  it  became  black. 

('97).  MIYOSHI,  MANABU.  Ueber  das  massenhafte 
Vorkommen  von  Eisenbacterien  in  den 
Thermen  von  Ikao.  Journ.  of  the  Coll.  of 
Science,  Imperial  Univ.,  Tokyo,  Japan,  vol. 
x,  Pt.  11,  1897,  pp.  139-142. 

('97).  MIGULA.    See  in. 

('97).  LAFAR.    See  in. 

('04).  SCHORLER,  B.  Beitrage  zur  kenntnis  der  Eisen- 
bakterien.  Centralb.  f.  Bakt.,  1904,  Bd.  xn, 
pp.  681-695. 


L.    Sulphur-Bacteria. 

C86).  CERTES  AND  GARRIGOU.    See  xxxiv. 

('87).  WINOGRADSKY,  SERGIUS.     Ueber  Schwefelbak- 

terien.     Bot.    Zeitung.    1887,    Bd.    XLV,   col. 

489,  513,  529,  545,  569,  585,  606.    3  figs. 
('88).  WINOGRADSKY,  S.     Sur  la  morphologic  et  la 

physiologic     des     sulfobacteries.       Beitr.     z. 

Morphol.  und  Physiol.  d.  Bakterien.,  fasc.  I. 

Leipzig,  1888. 
Not  seen. 

('89).  WINOGRADSKY,  S.  Recherches  sur  les  sulfo- 
bacteries. Ann.  de  1'Inst.  Pasteur,  T.  in, 
1889,  pp.  49-60. 

('89).  DE  TONI  AND  TREVISAN.  Sulphur  bacteria.  See 
Saccardo's  Sylloge  Fungorum,  vol.  8,  p. 
1,027.  Species  granula  sulphuris  secernentes. 

('93).  ZELINSKY,  N.  D.  Ueber  Schwefelwasserstoff- 
garung  im  Schwarzen  Meere  und  den 
Limans  von  Odessa.  Fortschr.  d.  russ. 
chem.  u.  phys.  Gesellsch.,  Bd.  xxv,  Part  V, 
1893,  PP-  298-303.  (In  Russian.) 

('95)-  YEGOUNOW,  M.  Sur  les  sulfo-bacteries  des 
limans  d'Odessa.  Archiv.  des  sci.  bio.  de 
1'Inst.  imperial  de  med.  exper.  de  St.  Peters- 
bourg,  vol.  in,  1895,  pp.  381-397.  Rev.  in 
Ann.  de  Micr.,  T.  vn,  1895,  pp.  281-282. 

('95).  BEYERINCK,  M.  W.  Ueber  Spirillum  desul- 
furicans  als  Ursaohe  von  Sulfatreduotion. 
Centralb.  f.  Bakt.,  2  Abt.,  Bd.  i,  1895,  pp. 
49-59  and  104-114. 

('97).  MIGULA.    See  in. 

('97).  LAFAR.    See  in. 

('97).  MIYOSHI,  MANABU.  Studien  iiber  Schwefel- 
rasenbildung  und  die  Schwefelbacterien  der 
Thermen  yon  Yirmoto  bei  Nikko.  Jour. 
College  Sci.,  Imp.  Univ.  Tokyo,  vol.  x,  Pt. 
11,  pp.  143-173,  1897,  i  plate.  Rev.  in  The 
American  Naturalist,  vol.  xxxn,  1898,  pp. 
456-457- 

(*0i).  CONN.    See  in. 


262 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('02).  NATHANSOHN,  ALEXANDER.  Ueber  eine  neue 
Gruppe  von  Schwefelbacterien  und  ihre 
Stoffwechsel.  Mitt.  a.  d.  zool.  Stat.  Neapel., 
Bd.  xv,  1902,  pp.  655-680. 

('03).  HINZE,  G.  Thiophysa  volutans,  ein  neues 
Schwefel-Bacterium.  Ber.  der  d.  hot. 
Gesellsch.,  Bd.  xxi,  Hft.  6,  July,  28,  1903, 
PP.  309-316. 

('03).  VAN  DELDEN.    See  xxiv. 


('79). 
('95). 

('95). 
('95). 

(95). 
(96). 
(•96). 
('96). 
('96). 
('96). 
('96). 
('96). 

('97). 
(97). 
(97). 
('97). 
('98). 


LI.     Bacteria  in  Prehistoric  Times. 

VAN  TIECHEM,  PH.    Sur  la  fermentation  buty- 

rique   (Bacillus  amylobacter)   a  1'epoque  de 

la  houille.    C.  R.  des  se.  de  1'Acad.  des  sci., 

Paris,  1879,  T.  LXXXIX,  pp.  1,102-1,104. 
RENAULT,  B.    Sur  quelques  bacteries  des  temps 

primaires.       Bull,     du     Museum     d'histoire 

naturelle,    Paris,   annee    1895,   T    i,   No.   4, 

pp.  168-172,  4  figs. 
RENAULT,    B.      Sur    quelques    Bacteries    due 

Dinantien  (Culm).     C.  R.  des  se.  de  1'Acad. 

des  Sci.,  Paris,  T.  cxx,  1895,  pp.  162-164. 
RENAULT,   B.     Sur  quelques  inicrococcus    du 

Stephanien,   terrain    houiller    superieur.      C. 

R.  des  se.  de  1'Acad.  des  sci.,  Paris,  T.  cxx, 

1895,  pp.  217-220. 
RENAULT,  B.    Sur  quelques  bacteries  anciennes. 

Bull,  du  Mus.  d'Hist.  nat,  Paris    1895,  T.  i, 

pp.  247-252,  6  figs. 
RENAULT,    B.      Sur    quelques    bacteries    de- 

voniennes.     C.  R.  des  se.  de  1'Acad.  des  sci., 

Paris,  T.  cxxn,  1896,  pp.  1,226-1,227. 
RENAULT.    Houille  et  Bacteriacees.    Soc.  d'nat. 

d'Autun,  Bull,  ix,  Autun,  1896,  pp.  475-500, 

i  pi. 
RENAULT,  B.     Les  bacteriacees  de  la  houille. 

C.  R.  des  se.  de  1'Acad.  des  sci.,  Paris,  T. 

cxxin,  1896,  pp.  953-955. 
RENAULT,  B.    Les  Bacteries  devoniennes  et  le 

genre  Aporoxylpn  d'Unger.     Bull,  du  Mus. 

d'Hist.  nat.,  Paris,  1896,  T.  n,  pp.  201-203. 
RENAULT,   B.     Notes    sur   quelques   nouvelles 

bacteries    fossiles.      Bull,    du    Mus.    d'Hist. 

nat.,  Paris,  1896,  T.  11,  pp.  285-288,  4  figs. 
RENAULT,  B.     Recherclies  sur  les  bacteriacees 

fossiles.     Ann.   des  sci.  nat.  hot.,  vin   serie, 

T.  ii,  1896,  pp.  275-349,  with  46  figures. 
RENAULT,  BERNARD.    Les  bacteries  devonienne 

et  le  genre  Aporoxylon  d'Unger.     Bull.  d.  1. 

Soc.  d'Hist.  nat.  d'Autun,  T.  ix,   1896,  pp. 

139-142,  of  the  Proces-verbaux  des  se. 
RENAULT,  B.     Les  bacteriacees  des  bogheads. 

C.  R.  des  se.  de  1'Acad.  des  sci.,  Paris,  T. 

cxxiv,  1897,  PP-  1,315-1,318. 
RENAULT,  B.    Les  bacteriacees  et  les  bogheads 

a  Pilas.     Bull.   d.   Mus.   d'Hist.  nat.,   Paris, 

T.  in,  1897,  pp.  33-39,  4  figs. 
RENAULT,  B.     Les  bacteriacees  des  bogheads. 

Bull.  d.  Mus.  d'Hist.  nat.,  Paris,  T.  in,  1897, 

pp.  251-258,  6  figs. 
RENAULT,  BERNARD.    Bogheads  et  bacteriacees. 

Soc.  d'histoire  naturelle  d'Autun.     x  Bulle- 
tin, 1897,  PP-  433-469,  18  text  figures. 
RENAULT,   B.     Les   microorganismes   des   lig- 
nites.     C.    R.    des    se.    de   1'Acad.    des    sci., 

Paris,  T.  cxxvi,  1898,  pp.  1,828-1,831. 


('98).  RENAULT,  B.,  ET  ROCHE,  A.  Du  mode  de 
propagation  des  bacteriacees  dans  les  com- 
bustibles fossiles  et  du  role  qu'elles  ont  joue 
dans  leur  formation.  Soc.  d'histoire  nat. 
d'Autun,  ix  Bull.,  1898,  pp.  133-147,  in  the 
Proces-Verbaux  d.  se. 

('oo).  LEMIERE,  L.  Transformation  des  vegetaux  en 
combustibles  fossiles.  Essai  sur  le  role  des 
ferments.  Congres  geologique  international 
de  1900,  Paris,  T.  I,  1901,  pp.  502-520. 


LII. 


Preparation  of  Slides,  Cultures,  Etc.,  for 
Museums,  &c. 


('80).  KAISER,  EDUARD.  Verfahren  zur  Herstellung 
einer  tadellosen  Glycerin-Gelatin.  Bot. 
Central!).,  Bd.  I,  1880,  pp.  25-26. 


('83).  GROVE,  W.  B.  New  methods  of  mounting  for 
the  microscope.  (Hillhouse's  method  for 
glycerine  mounting.)  Midland  Naturalist, 
vol.  vi,  1883,  p.  166.  Journal  of  the  R. 
Microscop.  Soc.,  London,  August,  1883,  p. 
599- 

Accordiugto  Hillhouse,  as  reviewed  by  Dippel  in  Botan. 
Centralb.,  p.  159,  Vol.  xvi,  1883,  glycerin  mounts  are 
readily  made  tight  by  substituting  Canada  balsam  dis- 
solved in  turpentine  for  ordinary  cements.  Ring  in  ordi- 
nary way.  Hillhouse  says  that  a  drop  of  glycerin  on 
glass  can"  be  covered  by  a  drop  of  balsam,  and  the  latter 
will  spread  over  it  and  adhere  firmly  to  glass  around  it 
on  all  sides,  inclosing  it  completely. 

('87).  SOYKA,  J.  Ueber  ein  Verfahren,  Dauerpr;ip:i- 
rate  von  Reinkulturen  auf  festem  Nahrboden 
herzustellen.  Centralb.  f.  Bakt.,  1887,  Bd.  i, 
PP.  542-544- 

('88).  JACOBI,  ED.  Hartung  und  Farbung  von  Plat- 
tenkulturen.  Centralb.  f.  Bakt.,  1888,  in 
Bd.,  pp.  536-538. 

('88).  SOYKA,  J.,  UND  KRAL,  F.  Vorschlage  und 
Anleitungen  zur  Anlegung  von  bacteriolo- 
gischen  Museen.  Zeitschr.  f.  Hyg.,  1888, 
Bd.  IV,  pp.  143-150. 

('89).  KRAL,  FRANZ.  Weitere  Vorschlage  und  An- 
leitungen zur  Anlegung  von  bakteriologis- 
chen  Museen.  Zeitschr.  f.  Hyg.,  Bd.  v,  1889, 
PP.  497-505. 

('89).  SCHILL.  Kleine  Beitrage  zur  bakteriologischen 
Technik.  i.  Konservirung  von  Flatten-  und 
Reagensglaskulturen.  6.  Schimmelpilze 
hindert  man  i.m  Waohsthum.  Centralb.  f. 
Bakt.,  1889,  v  Bd.,  Marz  i,  No.  10,  pp.  337- 
340. 

Cultures  are  covered  for  24  hours  with  a  fluid  consisting 
of  equal  parts  of  alcohol  and  glycerin,  to  which  has 
been  added  i  part  per  :oo  of  a  I  per  cent  solution  of  mer- 
curic chloride.  Preparations  treated  in  this  way  are 
said  to  remain  unchanged  for  years. 

Camphor  is  said  to  hinder  thegrowth  of  molds  without 
interfering  seriously  with  bacteria. 

('92).  DAWSON,  CHARLES  F.  Eine  Methode,  Dauer- 
kulturen  von  Bakterien  hermetisch  zu  ver- 
schliessen.  Centralb.  f.  Bakt.,  xn  Bd.,  1892, 
pp.  720-721. 


STOCK-CULTURES,  ETC.;  COLOR-CHARTS;    PHOTOGRAPHY,    ETC. 


263 


('93)-  HAUSER,  G.  Ueber  Verwendung  des  For- 
malins zur  conservirung  von  Bacteriencul- 
turen.  Miinchen.  nied.  Wochenschr.,  1893. 
Bd.  xr.,  pp.  567-568.  Rev.  in  Centralb.  f. 
Bakt,  Bd.  xiv,  1893,  p.  290. 

Ten  or  fifteen  drops  of  fresh  formalin  are  put  on  filter 
paper  and  placed  tinder  the  cover  of  the  Petri-dish  cul- 
ture. This  is  then  exposed  to  the  vapor  of  formalin  in  a 
close  room  lined  with  wet  filter  paper,  15  drops  of  forma- 
lin being1  placed  on  cotton  and  introduced  for  each  1000 
cc.  of  air  space.  As  the  formalin  penetrates  the  deeper 
layers  of  gelatin  only  slowly,  a  thin  layer  should  be  used 
for  liquefying:  organisms.  The  formalin  should  be 
allowed  to  act  for  several  days,  and  be  renewed  once  or 
twice.  The  gelatin  appears  to  be  permanently  disin- 
fected, will  not  melt  at  any  temperature,  and  is  un- 
changed in  appearance.  To  have  permanent  preparations 
it  is  only  necessary  to  keep  them  from  drying  out. 

fy.3).  HAUSER,  G.  Weitere  Mitteilungen  iiber  Ver- 
wendung  des  Formalins  zur  conservirung 
von  Bacterienculturen.  Miinchen.  med. 
Wochenschr.,  1893,  Bd.  XL,  pp.  655-656,  No. 
35-  Rev.  in  Centralb.  f.  Bakt.,  Bd.  xiv,  1893, 
pp.  468-469. 

Describes  a  method  of  fixing  and  mounting  colonies 
taken  from  gelatin  plate  cultures. 

('94).  KRUECKMANN.  EMU..  Eine  Methode  zur  Her- 
stellmig  bakteriologischer  Mttseen  ttnd  Kon- 
servierung  von  Bakterien.  Centralb.  f. 
Bakt,  Bd.  xv,  1894,  pp.  851-857. 

Fixes  cultures  with  mercuric  chloride,  etc.,  and  pre- 
serves them  in  formalin,  air-tight,  in  the  dark. 

('97).  FAKES,  W.  C.  C.,  AND  EYRE,  J.  W.  Formalin 
as  a  preservative  for  cultivations  of  bac- 
teria. Jour,  of  Path,  and  Bact.,  vol  iv,  1897, 
pp.  418-420.  Also  a  separate,  3  pp. 

('oi).  CONN,  H.  W.  How  can  bacteria  be  satisfac- 
torily preserved  for  museum  specimens? 
Science,  n.  s.,  vol.  xin,  1901,  p.  326. 


LIII.   Stock-Cultures,  How  Best  Kept;   Vitality  on 
Media. 

('89).  CZAPI.EWSKI,  E.  Zur  Anlage  bakteriologischer 
Museen.  Centralb.  f.  Bakt.,  vi  Bd.',  1889, 
pp.  409-411. 

This  method  consists  in  limiting  the  amount  of  air 
which  can  reach  the  culture  by  saturating  the  upper  part 
of  the  cotton  plug  with  melted  paraffin.  The  chief  objec- 
tion to  it  is  the  increased  difficulty  of  cleaning  the  dis- 
carded tubes. 

('98).  LUNT.  On  a  convenient  method  of  preserving 
living  pure  cultures  of  water  bacteria.  Rev. 
in  Centralb.  f.  Bakt.,  xxm  Bd.,  1898,  pp. 
795-796. 

Certain  water-bacteria  may  be  kept  alive  for  two  years 
or  more  in  sterile  water,  i.  e.  much  longer  than  in  ordi- 
nary culture-media. 

('oo).  BOLLKY,  HENRY  L.  The  duration  of  bacterial 
existence  and  [in  ?]  trial  environments. 
Centralb.  f.  Bakt.,  2  Abt,  vi  Bd.,  1900,  pp. 
33-38. 

Reports  getting  a  good  growth  of  Bacillus  amylovorns 
and  Bact.  dianthi  in  agar  and  bouillon  by  transfers  from 
cultures  which  had  been  hermetically  sealed  for  9  years. 
Tests  of  the  pathogenic  power  of  these  cultures  appear 
not  to  have  been  made. 

('oi).  SCHUI.TZ.    See  vi. 


LIV.    Color-Charts ;  Nomenclature  of  Colors. 

('86).  RIDGWAY,  ROBERT.  Nomenclature  of  colors 
for  naturalists.  195  water  colors  on  ten 
plates,  with  rules  for  making  the  same  and 
a  general  discussion  of  colors.  Boston,  Lit- 
tle, Brown  &  Co.,  1886. 

Valuable,  but  out  of  print.  Especially  useful  on 
account  of  the  number  of  colors.  Another  edition  in 
preparation. 

('94).  SACCARDO,  P.  A.  Chromotaxia  sen  nomen- 
clator  colorum  polyglottus  additis  speci- 
minibus  coloratis  ad  usum  botanicorum  et 
zoolpgortim.  2d  ed.  Padua.  Typis  Semi- 
narii,  1894,  8vo.,  22  pp.,  with  two  tables  con- 
taining 50  colors. 
A  cheap  and  useful  color  scheme  for  botanists. 

('95).  Color  chart  under  Spectrum,  in  the  Standard 
Dictionary,  Funk  and  Wagnalls,  New  York, 
1895- 
This  may  be  had  separately. 

('95).  SHUTTLEWORTH,  E.  B.  Nomenclature  of  col- 
ors for  bacteriologists.  Jour.  Am.  Pub. 
Health  Asso.,  Oct.,  1895,  Annual  vol.  xx, 
pp.  403-407. 

('98).  PRANG,  Louis.  The  Prang  standard  of  color. 
Popular  ed.,  Boston,  1898.  Folio. 

C — )•  Chart  of  Spectrum  Scales  made  from  the 
Bradley  colored  papers.  Milton  Bradley 
Co.,  Springfield,  Mass.  A  small  sheet  (about 
3/4x754  inches)  with  90  colors.  Also  a 
large  folded  chart  (11x28  indies). 

Colors  are  bright,  but  must  be  carefully  protected  from 
the  light. 


LV.    Photography  and  Photomicrography. 

C/7).  KOCH,  ROBERT.  Verfahren  zur  Untersuchun- 
gen,  zum  Conserviren  und  Photographiren 
der  Bacterien.  Cohn's  Beitrage,  II  Bd.,  3 
Heft,  Breslau,  1877,  pp.  399-434,  with  24 
photomicrographs  on  3  plates. 

('81).  KOCH,  R.  Zur  Untersuchung  von  pathogenen 
Organismen.  Mitth.  aus  dem  Kais.  Gesund- 
heitsamte,  Bd.  I,  1881,  pp.  1-48. 

The  paper  is  illustrated  by  84  heliotypes  from  photomi- 
crographs. 

('83).  STERNBERG,  GEO.  M.  Photomicrographs  and 
how  to  make  them ;  pp.  xv,  204,  with  twenty 
plates  of  photomicrographs.  James  R.  Os- 
good  &  Co.,  Boston,  1883. 

('87).  CROOKSHANK,  EDGAR  M.  Photography  of  bac- 
teria. Illustrated  with  86  photographs  re- 
produced in  autotype,  pp.  xix,  64,  London, 
H.  K.  Lewis,  1887. 

('87).  Roux,  E.  La  photographic  appliance  a  1'etude 
des  microbes.  Ann.  de  1'Inst.  Pasteur,  T.  I, 
1887,  pp.  209-225. 

("88).  ZETTNOW,  E.  Das  Kupfer-Chrom-Filter. 
Centralb.  f.  Bakt..  1888.  Bd.  iv,  pp.  51-52. 

This  light  filter  is  well  adapted  to  photographing  bac- 
teria, whether  they  are  stained  red,  blue  or  violet. 

This  filter  is  made  as  follows:  For  use  with  sunlight, 
160  grams  copper  nitrate  and  14  grams  pure  chromic  acid 
diluted  with  water  to  250  cc.  More  convenient  to  prepare 
and  suitable  for  most  purposes  in  a  layer  l  to  1  cm.  thick 
is  175  grams  sulphate  of  copper  and  17  grams  bichromate 
of  potash  dissolved  in  I  litre  of  water.  Thecopper-chrom- 
filter  transmits  only  a  small  portion  of  the  spectrum , 
viz.,  those  yellow-green  rays  which  act  most  strongly 
upon  erythrosin  plates.  For  the  concentrated  solution 
these  rays  are  from  wave  length  580  to  560  :  more  diluted 
from  590  to  545. 


264 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


('88).  NEUHAUSS,  R.  Verschiedenes  uber  Micro- 
photographie.  Zeitschr.  f.  Mikr.,  Bd.  v,  pp. 
484-486,  1888. 

('90).  NEUHAUSS,  RICHARD.  Lehrbucb  der  Mikro- 
photographie.  pp.  xi,  273,  with  61  wood 
cuts,  4  autotypes,  2  collotype  plates,  and  I 
photogravure.  Braunschweig,  Harald  Bruhn, 
1890. 
A  very  useful  book. 

("90).  PRINGLE,  ANDREW.  Practical  photo-micro- 
graphy; by  the  latest  methods.  The  Scovill 
&  Adams  Co.,  New  York,  1890,  pp.  183,  ix, 
42  figs.  Frontispiece  and  6  plates. 

('90  to  '99).  VOGEL,  H.  W.  Handbuch  der  Photo- 
graphic. Vier  Theile  enthaltend  die  photo- 
graphische  Chemie,  Optik,  Praxis  und 
Kunstlehre.  Berlin.  I  Thiel.,  1890,  pp.  XVI, 
351,  12  plates;  n  Theil.,  1894,  pp.  xi,  367; 
in  Theil.,  Abt.  I,  1897,  pp.  x,  311,  Abt.  II, 
1899,  PP-  x,  159. 

('99).  The  photo-miniature.  Tennant  and  Ward, 
New  York.  Begun  in  1899. 

A  series  of  small,  inexpensive  volumes,  by  various 
autliors,  on  various  phases  of  photography.  Some  of 
them  excellent.  About  70  numbers  up  to  beginning  of 
1905- 

('99).  HUBBARD,  J.  G.  Color  screens  as  applied  to 
photomicrography.  Jour.  Bost.  Soc.  Med. 
Sci.,  vol.  in,  1899,  pp.  297-301. 

("99).  WRIGHT,  JAMES  H.  Examples  of  the  applica- 
tion of  color  screens  to  photomicrography. 
Jour.  Bost.  Soc.  Med.  Sci.,  vol.  in,  1899, 
pp.  302-307. 

('01).  ABNEY,  SIR  WILLIAM  DE  WIVELESLIE.  A 
treatise  on  photography.  Tenth  ed.  thor- 
oughly revised,  with  134  illustrations.  Long- 
mans, Green  &  Co.,  39  Paternoster  Row, 
London,  New  York,  and  Bombay,  1901,  pp. 
xvii,  425. 

An  excellent  book,  butsubjects  treated  very  briefly. 

('02).  WALMSLEY,  W.  H.  The  A  B  C  of  photo- 
micrography, pp.  viii,  155.  Tennant  and 
Ward,  New  York,  1902. 


LVI.    Methods  and  Systems  of  Classification. 

(See  also  III  and  X.) 

('38).  EHRENBERG.    See  v. 

('41).  DUJARDIN,  F£LIX.  Histoire  naturelle  des  zo- 
ophytes, infusoires  comprenant  la  physiolo- 
gie  et  la  classification.  Paris,  1841. 

('65).  DAVAINE.    See  v. 

C°5-'67).  TRECUL,  A.  Urocephalum.  C.  R.  des  se. 
de  1'Acad.  des  sci.,  1865,  T.  LXI,  p.  156  and 
432.  Ibid.  1867,  T.  LXV,  p.  513. 

The  form  of  bacterium  with  a  spore  at  one  end,  which 
is  swollen,  was  called  Urocephalum  by  Trecul. 

('72).  COHN.    See  v. 

('79).  TREVISAN.  Introduzzione  allo  studio  die  bac- 
teri.  Atti  d.  Inst.  Lombardo,  1879. 

('80).  WINTER.    See  in. 

('81).  ZOPF,  W.  Ueber  den  genetischen  Zusammen- 
hang  von  Spaltpilzformen.  Monatsbericht 
d.  Konigl.  preuss.  Akad.  d.  Wissenschaften, 
Berlin,  1881,  pp.  277-284,  i  plate.  See  also 
various  editions  of  Zopf  s  "Spaltpilze." 

('85)    DE  BARY.    See  in. 


('85).  KUENTSLER,  J.  De  la  position  systematique 
des  bacteriacees.  Jour,  de  micr.,  T.  ix,  1885, 
pp.  295-307. 

According  to  this  author  the  bacteria  are  of  animal 
origin,  the  nearest  relatives  being  the  Flagellata,  espe- 
cially the  astomous  forms.  There  are  transition  forms. 

('86?).  SCHROETER.    See  in. 

('86).  HUEPPE,  FERDINAND.  Die  Formen  der  Bak- 
terien  und  ihre  Beziehungen  zu  den  Gattun- 
gen  und  Arten.  With  24  wood-cuts.  Wies- 
baden, C.  W.  Kreidel's  Verlag,  1886,  pp. 
vin,  152. 

('89).  DE  TONI  AND  TREVISAN.  Sylloge  Schizomy- 
cetarum.  Forms  a  portion  of  vol.  vin  of 
Saccardo's  Sylloge  Fungorum.  Padua,  1889, 
pp.  923-1,087. 

Those  volumes  of  Saccardo's  Sylloge  Fuugorum  which 
are  out  of  print  have  been  reproduced  in  fac-simile 
(zincography)  by  R.  Friedlander  and  Sohn,  Berlin. 

('90).  MESSEA,  AL.  Contribuzione  allo  studio  delle 
ciglia  dei  batterii  e  proposta  di  una  classifica- 
zione.  Revista  d'igiene  e  Sanita  Pubblica, 
Anno  I,  1890,  pp.  513-528,  I  plate.  Bibliog- 
raphy of  19  titles.  Rev.  in  Centralb.  f.  Bakt., 
etc.,  Bd.  ix,  1891,  pp.  106-107,  and  in  Baum- 
garten's  Jahresbericht,  Bd.  vn,  p.  344. 

The  bacteria  are  classified  as  Gymnobacteria  and  Trich- 
obacteria.  The  latter  are  subdivided  into  four  groups  : 
Monotricha  (one  polar  flagellum),  Lophotricha  (a  tuft 
at  one  pole),  Amphitricha  (one  flagellum  at  each  end), 
and  Peritricha  (flagella  from  various  parts  of  the  body). 
These  names  are  not  used  in  a  generic  sense. 

('92).  WARD,  H.  MARSHALL.  On  the  characters  or 
marks  employed  for  classifying  the  Schizo- 
mycetes.  Annals  of  Botany,  vol.  6,  1892,  p. 
103- 

('94).  MIGULA,  W.  Ueber  ein  neues  System  der  Bak- 
terien.  Arbeiten  aus  dem  Bakt.  Institut  der 
Technische  Hochschule  zu  Karlsruhe,  Bd.  I, 
Hft.  2,  1894,  pp.  235-238. 

('95).  MIGULA,  W.  Schizomycetes.  Engler  and 
Prantl's  Die  Naturlichen  Pflanzenfamilien, 
Leipzig,  Wilhelm  Engelmann,  1895. 

('95).  FISCHER,  ALFRED.  Untersuchungen  iiber  Bak- 
terien.  Jahrb.  f.  wissensoh.  Botanik,  Bd. 
xxvn,  1895,  Hft.  i,  pp.  1-163.  5  plates. 

('96).  MEz,  CARL.  Der  heutige  Stand  der  bak- 
teriologischen  Systematik.  Botanisches 
Centralb.,  Bd.  LXVIII,  1806,  pp.  203-211. 

('97).  CHESTER,  FREDERICK  D.  A  preliminary  arrange- 
ment of  the  species  of  the  genus  Bacterium. 
Oth  Ann.  Rept.  of  the  Delaware  Coll.  Agr. 
Exp.  Sta.,  1897.  Also  a  separate,  pp.  93. 

('97).  MIGULA.    See  in. 

('97).  FISCHER.    See  in. 

Coo).  MIGULA.    See  111. 

('02).  THAXTER.    See  x. 

("03).  KENDALL,  ARTHUR  I.  A  proposed  classifica- 
tion and  method  of  graphical  tabulation  of 
the  characters  of  bacteria.  Proc.  Amer.  Pub. 
Health  Asso.,  Thirtieth  annual  meeting,  held 
at  New  Orleans,  La.,  Dec.,  1902,  vol.  xxvin, 
pp.  481-493.  Also  a  separate,  pp.  3-15.  Pub. 
1003. 

('03).  FORD,  WILLIAM  W.  The  classification  and  dis- 
tribution of  the  intestinal  bacteria  in  man. 
Studies  from  the  Royal  Victoria  Hospital, 
Montreal,  vol.  i,  No.  5  (Pathology  n), 
1003,  PP-  3-95.  3  tables.  Also  a  separate. 


CATALOGUES. 


265 


('03).  GAGE,  STEPHEN  DE  M,  AND  PHELPS,  EARLE  B 
On  the  classification  and  identification  o< 
bacteria,  with  a  description  of  the  card  sys- 
tem in  use  at  the  Lawrence  Exp.  Station  for 
Records  of  Species  (Lawrence,  Mass.). 
Proceedings  of  thirtieth  annual  meeting-  of 
American  Public  Health  Assn.,  New  Or- 
leans, La.,  Dec.,  1902,  vol.  xxvm,  pp.  494- 
505.  Pub.  Columbus,  O.,  1903.  Also  a 
separate,  pp.  12-23. 

('04).  PERKINS,  ROGER  G.  Bacillus  Mucosus  Capsu- 
latus.  A  study  of  the  group  and  an  attempt 
at  classification  of  the  varieties  described. 
Jour,  of  Infectious  Diseases,  vol.  I,  No.  i, 
1904,  pp.  241-267.  Also  a  separate. 

('05).  WINSLOW,  C.  E.  A.,  AND  ROGERS,  ANNE  P.  A 
revision  of  the  Coccaceae.  Preliminary 
communication.  From  the  Biological  labo- 
ratories of  the  Massachusetts  Institute  of 
Technology.  Science,  n.  s.,  Vol.  xxi,  1905, 
pp.  669-672. 

The  445  described  forms  are  reduced  to  31  types.  Five 
genera  are  recognized,  viz,  Diplococcus  and  Streptococ- 
cus, belonging  to  the  sub-family  Paracoccaceae,  and  Mi- 
crococcus,  Sarcina  and  Ascococcus,  belonging  to  the  sub- 
family Metacoccaceae. 


LVII.     Useful  Catalogues. 

Catalogues   and   addresses   of  instrument   makers, 
manufacturers  of  chemicals,  etc. : 
CARI,  ZEISS,  Jena. 

(1)  Microscopes  and  microscopical  accessories,  32 

ed.,  1902. 

(2)  Photographic  objectives  and  photo-optical  ap- 

pliances, 1901. 

(3)  Catalogue  of  instruments  and  appliances   for 

projection  and  photomicrography,  fourth  ed., 
1899. 

ERNST  LEITZ,  Wetzlar. 

Microscopes  and  accessory  apparatus.    Cat.  39.    U. 
S.  Branch:     Wm.  Krafft,  80  East  i8th  st,  near 
Broadway,  New  York. 
BAUSCH  AND  LOME,  Rochester,  N.  Y. 

(1)  Optical    apparatus,   microscopes,   photographic 

lenses. 

(2)  Chemical  apparatus,  bacteriological  apparatus. 
EIMER  AND  AMEND,  New  York. 

Chemical  and  physical  apparatus,  1903.    Am.  agents 
for  Zeiss. 

WHITALL  AND  TATUM,  Philadelphia,  New  York,  and 
Boston. 

Glassware. 
F.  AND  M.  LAUTENSCHLAEGER,  Berlin. 

Cat.  No.  60.    Bacteriologie,  Chemie,  Asepsis. 
MAISON  WIESNEG  (P.  Lequeux),  64  Rue  Gay-Lussac, 
Paris. 

Catalogue     des     appareils     de     bacteriologie     et 

d'hygiene. 
M.  SCHANZE,  Leipzig. 

Preisverzeichnis  von  Mikrotome. 
DR.  HERMANN  ROHRBECK,  Berlin. 

Brood-ovens,  thermo-regulators,  etc. 
DR.  G.  GRUEBLER  AND  Co.,  Leipzig. 

Preislisten  von  Farbstoffe  und  Reagentien. 


Griibler's  stains  may  be  obtained  in  the  U.  S.  from 
Wm.  Krafft,  80  East  i8th  st.,  near  Broadway, 
New  York. 

Koenigliohe  Pprzellan-Manufactur,  Berlin. 

Preis-Verzeichniss,  No.  v.  Gerathschaften  zu 
chemischen  und  pharmaceutischen  Zwecken.  i 
Jan.,  1899. 

Price  list  of  chemical  apparatus  manufactured  and 
sold  by  C.  Gerhardt,  Bonn  am  Rhein,  Ger- 
many. 

P.  J.  KIPP  u.  ZOHNEN,  Delft,  Netherlands.     Makers 

of  the  Reinhold-Giltay  microtome. 
DR.  ROB.  MUENCKE. 

Catalog  fiber  chemische  Apparate  und  Gerathschaf- 
ten, 1900,  Berlin,  N.  W.,  Luisen-Strasse  58. 
Preis  5  mark.  pp.  600. 

DR.  PETERS  &  ROST. 

(1)  Preis-Liste  fiber  Apparate  und  Utensilien  fur 

elektrochemische  und  elektrolytische  Ar- 
beiten.  Liste  No.  29.  Berlin,  1900,  pp.  48. 

(2)  Preis-Liste  fiber  Apparate  und  Utensilien  fur 

Bakterioloeie,  Hygiene,  Mikroscopie.  Rea- 
gentien, Farbstoffe,  Nahrboden,  Reincul- 
turen,  mikroscopische  Praparate.  Liste  No. 
30.  Berlin,  1900,  pp.  123. 

(3)  Preis-Liste  fiber  chemische  Apparate  und  Uten- 

silien fur  wissenschaftliche  und  Fahriks- 
Laboratorien,  Chemicalien,  Reagentien, 
Normal-Losungen.  Liste  No.  28.  Berlin, 
1002,  pp.  534. 

(4)  Preis-Liste  fiber  physikalische  Apparate.    Liste 

No.  37.    Berlin,  1902,  pp.  iv,  383. 

MAX  KAEHLER  AND  MARTINI,  Berlin. 
Chemical  and  Bacteriological  Apparatus. 

The  above  five  catalogues  or  their  equivalent  may 
now  be  obtained  from  the  following: 

Vereinigte  Fabriken  ffir  Laboratoriumsbedarf,  ges. 
M.  B.  H.,  ChauseestT.  3.  Berlin.  N.,  formerly  Max 
Kaehler  &  Martini.  Dr.  Peters  &  Rost.  New  York 
Office :  Laboratory  and  School  Supply  Co.,  20-24 
East  20th  street,  New  York. 

EMU,  GRF.INER,  New  York. 

Bacteriological  apparatus,  glassware,  high-grade 
glass  blowing.  Very  reliable. 

GREINER  &  FRIEDRICHS,  Stutzerbach,  Germany. 

Glassware. 
C.  P.  GOERZ. 

Price  list  of  double-anastigmat  lenses  and  of  the 
Goerz-Anschutz  camera.  Berlin-Friedenau,  and 
52  East  Union  Square,  New  York. 

FOI.MER  AND  SCHWING,  407  Broome  street,  New  York. 
High-grade  cameras. 

HENRY  HEIL  CHEMICAL  Co.,  298-312  S.  Fourth  street, 
St.  Louis,  Mo. 

Illustrated  catalogue  and  price  list  of  chemical  and 
physical  apparatus  and  instruments  for  labora- 
tories, chemists,  iron  and  steel  works,  smelters, 
assayers,  mines,  sugar  refineries,  schools,  col- 
leges, universities,  etc. 

SPENCER  LENS  Co.,  Buffalo,  N.  Y. 
Microscopes.     Excellent     lenses,    especially     low- 
power  apochromatics  and  a  new  2  mm.  oil  im- 
mersion achromatic. 


266 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


BECKER,  CHRISTIAN. 

Price  list  of  balances  and  weights  of  precision. 
Factory :  New  Rochelle,  N.  Y. ;  New  York  office : 
7  Maiden  Lane. 

THE  KNY-SCHEERER  Co.,  225-233  Fourth  avenue,  New 
York. 

(1)  Illustrated  Catalogue  of  Surgical  Instruments, 

1902,  pp.  LVI,  and  pp.  1,001-1,061,  2,001-2,140, 
3,001-3,192,  4,001-4,102,  and  5,001-5,180. 

(2)  Aseptic  Surgical  Furniture  and  General  Hos- 

pital  Supplies,  6   ed.,    1902,   pp.  232.     Fully 
illustrated. 

E.  H.  SARGENT,  &  Co.,  143  and  145  Lake  street,  Chi- 
cago. 

Importers    and    makers     of    laboratory     supplies, 

price    list    of    apparatus,     chemicals,     reagents, 

tissue-stains,    microscopical    and    bacteriological 

supplies,  etc. 

JAMES  T.  DOUGHERTY,  409  and  411  W.  Fifty-ninth 

street,  New  York. 

Scientific  apparatus.  Sole  United  States  agent  for 
Carl  Reichert,  Vienna,  Austria,  microscopes, 
microtomes,  and  polarizing  apparatus. 

THE     VblGTLAENDER     &     S*ON     OPTICAL     Co.,     137     W. 

Twenty-third  street,  New  York. 
Collinear  and  telephoto  lenses,  porro  prism  binoc- 
ulars, etc. 
WARNER  &  SWASEY,  Cleveland,  Ohio. 

Excellent  binocular  field  glasses. 
CHARLES  J.  Ross  Co.,  1525  Fairmount  avenue,  Phila- 
delphia, Pa. 

Excellent  quality  of  drawing-board,  heavily  coated, 
so  as  to  permit  of  any  number  of  erasures.  Rec- 
ommended by  Dr.  Roland  Tfaaxter  and  by  the 
•writer. 

THE  CENTURY  CAMERA  Co.,  Rochester,  N.  Y. 
Excellent    cameras    embodying   many    convenient, 

time-saving  devices. 
STANLEY  PHOTOGRAPHIC  DRY  PLATE  Co. 

Cheap  dry-plates  of  good  quality. 
M.  A.  SEED  DRY  PLATE  Co.,  St.  Louis,  Mo. 
Standard  dry-plates.     Non-curling  films.     Isochro- 
matic,  ordinary,  and  non-halation  plates  of  good 
quality. 

HAMMER  DRY  PLATE  Co.,  St.  Louis,  Mo. 
Both  Seed  and  Hammer  publish  interesting  little 
free  handbooks  on  negative  making,  etc. 


G.  CRAMER  DRY  PLATE  Co.,  St.   Louis,  Mo.     New 

York  Depot :    32  East  loth  street. 
CADETT  SPECTRUM  DRY  PLATE  Co.,  London,  England. 
THE  ILFORD  CHROMATIC  PLATE  Co.,  London,  England. 
The  3^4  by  4^4  plates  are  highly  recommended  by 
Dr.  Jeffrey  for  photomicrographic  work.     They 
cost  20  cents  per  dozen,  exclusive  of  duty. 

THE  EASTMAN  KODAK  Co.,  Rochester,  N.  Y. 
Roll  and  cut  films,  Kodoid  films,  non-curling  films, 
solio  paper,  velox  paper,  etc. 

WILLIS  AND  CLEMENTS,  Philadelphia,  Pa. 

Platinotype  paper. 
DR.  J.  C.  MILLER,  Denver,  Colo. 

Blue-print  papers  of  high  grade,  known  as  "French 
Satin,  Jr." 

G.  GENNERT,  24  and  26  East  ijth  street,  New  York. 
Hauff's  Ortol  developer,  white  glass  for  lantern- 
slide  covers,  etc. 

MALLINCKRODT  CHEMICAL  WORKS,  St.  Louis,  Mo.,  and 

New  York,  N.  Y. 

Photographic  chemicals  of  a  high  grade. 
KEUFFEL  &  ESSER  Co.,  New  York. 

127  Fulton  street  and  42  Ann  street.  Catalogue 
and  price  list  of  drawing  materials  and  surveying 
instruments.  Drawing  paper  of  any  quality  de- 
sired converted  into  blue-print  paper.  Branches 
in  Chicago,  St.  Louis,  and  San  Francisco. 

E.    SCHERING,   manufacturing   chemist,    Berlin,   Ger- 
many. 
KRAL'S  LABORATORY,  Prag,  Austro-Hungary. 

Der  gegenwartige  Bestand  der  Kral'schen  Samm- 
lung  von  Mikroorganismen,  Oct.,  1902.  Krai's  Bac- 
teriologisches  Laboratorium,  Prag,  I.  Kleiner  Ring 
11.  Telegramm-Addresse :  Krai's  Laboratorium. 

Cultures  of  several  hundred  sorts  of  bacteria  and 
of  some  fungi  may  be  had  from  this  laboratory. 
Authors  are  urged  to  send  their  new  species  to  Krai. 

W.  P.  Stender,  Leipzig,  Gerichtsweg  10.  Fabrik 
ttnd  Lager  von  Glasgegenstaenden  zur  Anfertigung 
mikroskopischer  Praeparate,  u.  s.  w. 

EAGLE  OXYGEN  Co. 

(See  p.  81.) 
DEFENDER  PHOTO  SUPPLY  Co.,  Rochester,  N.  Y. 

Argo  and  other  photographic  papers. 


ADDEN  DA. 


Page  46,  paragraph  b,  for  "  fat "  read  casein.  The  fluid  has  a  soapy  feeling,  but 
"  saponification  "  also  is  probably  not  the  proper  term.  What  actually  occurs  is  a 
matter  for  the  chemist  to  determine.  The  inoculated  milk  shows  no  change  at  first, 
but  gradually  becomes  intensely  alkaline  and  clears  synchronously,  without  coagula- 
tion or  precipitation.  By  addition  of  acids,  or  concentrated  solutions  of  sodium 
chloride,  copper  sulphate,  etc.,  the  transparent  fluid  may  now  be  filled  with  a  white 
flocculence,  which  slowly  settles  to  a  bulky  caseous  precipitate,  leaving  a  superna- 
tant clear,  pale  whey.  It  usually  requires  a  month  or  six  weeks  for  the  inoculated 
milk  to  become  entirely  transparent,  but  a  similar  transparency  may  be  produced  at 
once  in  check  tubes  of  milk  by  adding  a  few  drops  of  ammonia-water. 

On  testing  cultures  grown  for  a  few  days  in  "  nitrate  bouillon,"  as  described 
on  p.  63,  it  happens  frequently  that  there  is  no  nitrite  reaction.  It  is  then  neces- 
sary to  know  whether  nitrate  is  actually  present  in  the  bouillon.  Usually  cultures 
of  Bacillus  coli,  or  some  other  known  nitrate-reducing  organism  serve  this  purpose. 
If  such  cultures  have  not  been  provided,  the  test  for  nitrates  may  be  made  with 
diphenylamin  dissolved  in  strong  sulphuric  acid.  On  addition  of  a  few  drops  of 
this  reagent  there  is  an  immediate,  evanescent,  deep-blue  reaction  if  nitrates  are 
present.  The  reagent  is  prepared  by  slowly  dropping  90  cc.  of  c.  p  sulphuric  acid 
into  10  cc.  of  pure  water,  after  which  i  gram  of  diphenylamin  is  added,  and  the 
solution  preserved  in  a  glass-stoppered  bottle. 

267 


I  NDEX. 


Page. 

Abbe  camera 130 

Abstracts,  making  of 114 

Acclimatization  to  high  temperatures 222 

Acetic  alcohol. 

as  a  fixing  fluid 202 

with  mercuric  chloride 202 

Acetone,  for  use  in  fermentation-tubes 52 

Acid-fast  bacteria 188 

Acid-forming  bacteria,  demonstration  by  plate 

cultures  51,233 

Acid  gelatin,  effect  on  growth 30 

Acid, 

hydrochloric,  in  culture-media 98 

oxalic,  actively  germicidal 249 

Acids, 

action  on  enzymes 68 

effect  on  bacteria 249 

formation  of,  in  milk-cultures 46 

method  of  estimating  in  plants,  when  not 

free  207 

spilled,  neutralization  of 107 

toleration  of,  by  bacteria 70, 249 

vegetable,  sensitiveness  of  bacteria  to 69 

Achromatium  162 

Acrolein,  germicidal  action  of 252 

Achromatic  objectives, 

disadvantages  of 140 

with  excellent  definition 265 

Ae'robe,  Pasteur's  definition  of 230 

Agar-agar, 

combined  with  bouillon  and  sugars 33,5 1 

commercial,  source  of 31, 225 

composition  of 32, 223 

cultures  on,  for  flagella  staining 20 

filtration  of 33, 224, 225 

glycerinated  50, 196 

litmus  lactose 196 

melting  point  of 32 

methods  of  making 223,  224, 225 

nutrient,  centrifuging  of 225 

nutrient,  temperature  at  which  poured 105 

per  cent  used  for  media 35 

rate  of  diffusion  in 224 

roll-cultures  of 36 

softening  of,  by  bacteria 32 

solidification  of 32 

source  of 31 

standard  nutrient,  preparation  of 33,195 

sterilization  of 98, 99 

variation  on,  of  same  organism 180 

with  neutral  red 230 

without  nutrients 229 


-       Page. 

Agar-block  method  for  study  of  antagonism. .     73 
Agar-plates,    for    detection    of    acid-forming 

colonies  51 

Agfa-intensifier  142 

Air, 

bacteria  infrequent  in  Arctic 255 

bacteria  rare  in  upper  strata  of 255 

of  mountains,  bacteria  rare  in 255 

over  the  sea,  bacteria  rare  in 255 

Air-currents,  danger  of 103, 106 

Air-shaft,  for  ventilation  of  dark-room 149 

Albumen,  egg, 

Mayer's  119 

preparation  of,  for  culture-medium 48 

Alcohol, 

as  a  fixing  fluid 8, 202 

•ethyl,  for  use  in  fermentation-tubes 52 

methyl,  for  use  in  fermentation-tubes 52 

weak  germicidal  action  of 252 

Alkalies 

action  on  enzymes 68 

formation  of,  in  cultures 61 

sensitiveness  of  bacteria  to 69 

toleration  of,  by  bacteria 70 

Alkaline  methylene  blue 188 

Amaryllis,  inoculated  with  B.  hyacinth! 66 

Amiclo-bacteria  175 

Amins,  test  for 64 

Ammonia-bacteria  175 

Ammonia, 

oxidation  of,  by  bacteria 64 

test  for 64 

Ammonium, 

citrate,  relation  to  pigment-production 65 

lactate,  relation  to  pigment-production 65 

oxalate,  solvent  of  middle  lamella 67 

salts,  oxidation  of,  by  bacteria 64 

salts,  sterilization  without  loss  of  ammonia.     52 
succinate,  relation  to  pigment-production. . .     65 

Amoebobacter 163 

Anaerobe,  Pasteur's  definition  of 230 

Anaerobes  51 

Arens'  method  of  cultivating 231 

Buchner's  method  for  growing 231 

Bulloch's  method  of  obtaining 228 

Heim's  method 231 

Hesse's  method 230 

Klein's  apparatus  for 232 

-     on  culture-media  in  hydrogen,  etc 57 

present  in  milk 46 

Wright's  methods  of  cultivating 228 

269 


2-JO 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


Page. 

Anilin-dyes, 

differential  diagnosis  with 230 

watery  solutions  of '87 

Analin-dyes   (basic), 

affinity  of  bacteria  for 27 

use  of,  in  vegetable  tissues 29 

Anilin  fuchsin '^7 

Anilin  gentian  violet :°7 

Anilin  methyl  violet ^7 

Anilin  stains,  alcoholic  solutions  of 187 

Anilin-water T°7 

Animal-fluids  for  culture-media, 

beef-broth  45 

blood-serum 48 

egg-albumen  4° 

egg-yolk  •     49 

litmus-milk  4»,  196 

milk  46 

rice  cooked  in  milk 48 

Anschiitz,  normal  thermometers 78 

Antagonistic  action  of  bacteria 73 

Anthrax  organism, 

asporogenous  222 

behavior  when  injected  into  plants 89 

branching  in 216 

generic  name  for !7' 

Anthrax  spores,  substratum  influences  forma- 
tion of 219 

Antiseptics, 

effect  on  form  of  bacteria 252 

kinds  of 74 

literature  on 250 

Aplanobacter,  species  included  under  genus. .   171 
Apochromatic  objectives, 

in  photomicrography 139 

recommended  for  bacteriological  work 130 

Spencer 14° 

Zeiss 130 

Apparatus  necessary  in  laboratory 95 

Apple  blight,  due  to  B.  amylovorus 202 

Arnold  steam-sterilizer 47 

Arthrospores,  disputed  existence  of. ..  21, 158,218 
Asparagin, 

in  fermentation-tubes 52 

relation  to  pigment-production 65 

Atkinson,  method  for  photographing  poured 

plates  134 

Aujeszky,  spore-stain 219 

Autoclave,  for  sterilization  of  culture-media.  84,  98 
Bacillus, 

Cohn's  genus 158 

Fischer's  genus 157 

in  Migula's  classification 160 

Bacillus  amylovorus, 

absence  of  pigment  in 65 

distributed  by  insects 215 

effect  of  direct  sunlight  on 72 

enters  plant  through  nectaries 92 

flagella  hard  to  stain ; 190 

hosts  of 87 

nitrates  not  reduced  by 113 

non-sporiferous  158 

useful   for  comparison 29 


Page. 

Bacillus  aroideae 115 

hosts  of 86 

soft  rot  due  to 115 

stain  produced  by 65 

thermal  relations  of 86 

variation   in  shape  of  colonies  at  different 

temperatures 180 

Bacillus  Biitchlii,  inner  structure  of 279 

Bacillus  carotovorus, 

soft  rot  due  to 103 

stain  produced  by 65 

thermal  relations  of 86 

tissues  occupied  by 5,  6 

Bacillus  coli, 

effect  of  formalin  on 229 

how  differentiated  from  B.  amylovorus.  .  51,  113 
method  of  differentiating  from  B.  typhosus.  229 

useful  for  comparison 29 

vitality  of 72 

Bacillus  denitrificans,  size  of 18 

Bacillus  hortulanus,  isolation  of,  by  heat.   106,211 
Bacillus  oleraceae,  maximum  temperature  for.     86 

Bacillus  oligocarbophilus 241 

Bacillus  prodigiosus, 

culture-media  for 238,  239 

for  study  of  production  of  pigment 64 

magnesium  sulphate  necessary  for  pigment.  238 
pigmented  and  non-pigmented  races  of. .  64,222, 

238 

reaction  of  color  to  acids  and  alkalies 65 

Bacillus     rosaceus     metalloides,     rosette-like 

groupings  218 

Bacillus  tracheiphilus, 

absence  of  pigment  in 65 

action  of  block-tin  on 97 

behavior  in  fermentation-tubes 53 

behavior  toward  acids  and  alkalies 70 

distributed  by  insects 178,  215 

effect  of  f rezing  on 82 

effect  of  sodium  chloride  on 70 

susceptibility  to  dry  air 71 

viscidity  of 20 

Bacillus  typhosus, 

action  of  copper  on 74,  97 

behavior  when  injected  into  living  plants.  . .     89 

effect  of  formalin  on 229 

Hiss'  special  medium  for  differentiating. .. .  230 

non-sporiferous   158 

Wurtz's  method  of  differentiating 229 

Bacteria, 

action  on  photographic  plate 242 

amount  of  acid  tolerated 70 

animal  origin  of 264 

blue  pigment  forming,  in  cheese,  etc 237 

branched  forms 177, 217,  218,  237 

effect  of  chemicals  on 74,  250 

effect  of  freezing 79, 246,  247, 258 

effect  of  gases  on 58, 231 

effect  of  heat  on 75, 246 

effect  of  ions  on 244 

effect  of  light  on 71,  243 

effect  of  pressure  on 245 

hanging-block  cultures 22 


INDEX. 


271 


Page. 

Bacteria — continued. 

hanging-drop  cultures 22 

in  bread,  baking  does  not  destroy  all 261 

in  milk 196,  260 

isolation  from  diseased  tissues,  methods  of.     n 

limits  of  size 18 

long  vitality  of 72,  214,  263 

monotonous  morphology  of 25 

move  against  a  stream 253 

non-nitrifying  associated  with  nitrifying.  . .  241 
oxidation  of  ammonia  and  ammonium  salts 

by   64 

photographing  in  stained  tissues 136,  140 

prehistoric  362 

presence    in    diseased    tissues,   how   deter- 
mined        10 

rapidity  of  movement 26 

rare  in  Arctic 255 

rare  in  mountain  air 255 

sensitiveness  to  acids 69 

sensitiveness  to  alkalies 69 

species  found  in  meat-extracts 260 

thermal  range  of 75,  246 

toleration  limit  for  sodium  chloride.  ..    .   70,252 

Bacterial  ash,  alkaline 42 

Bacterial  disease  communicated  by  beetles....  92, 

178,215 

Bacterial  soft  rots,  similarity  of 8 

Bacteriology, 

milestones  in  progress  of 151 

prominent  workers  in 152 

Bacteriosis  of  walnut 174,  1/6 

prevention  of 93 

Bacterium, 

Cohn's  genus 166 

Ehrenberg's  genus 165 

iron  stored  by 261 

plant  parasites  belonging  to  genus 171 

Migula's  use  of  the  word 160, 165 

substituted  for  Pseudomonas 166,  171 

triloculare   165, 169 

Bacterium  campestre, 

communicated  by  slugs,  larvae,  etc 215 

effect  of  desiccation  on 71 

enters  plant  through  water-pores.  92,94,  102,  124 

enzymes  produced  by 68 

relation  to  soft  white  rots 73 

tissues  occupied  by 7,  10,  n,  12 

useful  for  comparison 29 

Bacterium  hyacinth!, 

effect  of  desiccation  on 71 

effect  on  Amaryllis  atamasco 66 

Bacterium  malvacearum, 

leaf  spots  due  to 95 

stomatal  infection 126 

Bacterium  pediculatum,  branching  of 221 

Bacterium  phaseoli, 

beans  infected  with,  by  way  of  stomata 92 

effect  of  direct  sunlight  on 72 

pigment  of 179 

Bacterium  pericarditidis, 

fiagellum  of 237 

green  fluorescence  of 65 


Page. 
Bacterium  pericarditidis — continued. 

viscidity  in  fluid  culture-media 42 

vitality  of 73 

Bacterium  solanacearum, 

distributed  by  insects 215 

optimum  temperature  for 86 

plants  inoculated  with 17,  202 

relation  to  soft  white  rots 73 

stain  produced  by 65 

Bacterium  syncyaneum,  blue  color  of,  in  acid 

milk   65 

Bacterium  syringae 64, 66,  135 

Bacterium  Stewart!, 

stock-cultures  of,  how  best  kept 72 

tissues  affected  by 4, 90 

Bacterium  termo, 

Cohn's  and  de  Bary's  account  of 166 

Dallinger  &  Drysdale's  conception  of 170 

flagella  of 20,  170, 219 

Bacterium  triloculare,  flagella  of 20,169 

Bacterium  vascularum, 

relation  to  red  stain  of  sugar-cane 66 

stock  cultures  of,  how  best  kept 72 

Bactridium,  as  a  genus  name 158 

Balsam,  for  mounting  sections 119 

Banti,  isolations  on  slant  media 227 

Barfoed's  reagent 208 

Bechamp,  views  concerning  morphology 176 

Beef-bouillon, 

proper  sterilization  of 98 

standard  peptonized 45,  195 

Beetles,  disease  spread  by 92, 178,  215 

Beggiatoa  162 

Bell-jars,  for  use  in  inoculations 108 

Benda's  iron-haematoxylin 188 

Benzoic  acid 74 

Beyerinck, 

agar    for    detection    of   acid-forming    colo- 
nies    51,  233 

agar  for  cultivation  of  nitrite  bacteria 199 

Bacillus  oligocarbophilus 241 

blue  pigment  bacteria  in  cheese 237 

fermentation  of  indigo 257 

Kapillarhebermikroscopirtropfenflasche 226 

root-tubercle  bacteria,  study  of 153 

thermotaxis  254 

Bilirubin  as  a  test  for  free  oxygen 57 

Billroth,  views  concerning  morphology 176 

Birds,  intestinal  tract  of,  bacteria  in 258 

Blood-serum  48 

Blue  prints,  making  of,  for  drawing  purposes.   150 
Bokorny,  amount  of  nutrients  needed  by  bac- 
teria    225 

Bolley,  on  longevity  of  bacteria 263 

Boni's  method  of  staining  capsules 194 

Bouillon, 

cultures  in,  for  flagella  staining 20,  190 

lead  acetate  in 51 

neutral  red  in 51 

of  varying  degrees  of  alkalinity  and  acidity.     51 

peptonized  standard 195 

salted   51 

sterilization  of 98, 99 


272 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Page. 

Bougie, 

Berkefeld's,  for  filtration  of  culture-fluids. .     43 
Chamberland's,    for    filtration    of    culture- 
fluids  43 

Chamberland's,    sterilization    of,    necessity 

for  frequent 44 

Bowhill's  flagella  stain I92 

Boxes  for  sterilization  of  pipettes 41 

Branched  forms 23, 177, 215, 216, 217, 218 

Bread, 

Bacteria  cause  stringiness  in 260,  261 

baking  does  not  fully  sterilize 261 

Bredig  und  Miiller,  enzymic  action  of  plati- 
num black 234 

Brenner,  black  rot  of  cabbage,  communicated 

by  aphides 215 

Bromide-prints  IS1 

Broomcorn, 

natural  infection  in 92,  150 

red  blotches  and  stripes,  relation  of  bacteria 

to  66 

Brownian  movement,  deceptive  nature  of. .   10,  26 
Brown  pigment,  bacteria  producing.   65,  21 1,  214,237 

Brunner  and  Zawadzki,  counting  plate 227 

Bubonic  plague,  nature  of  organism  causing..  214 
Buchner, 

effect  of  light  on  bacteria 244 

method  of  growing  anaerobes 231 

Bujwid, 

cholera  red 229 

comparison  of  Chamberland  and  Berkefeld 

filters  227 

Bunge's  flagella  stain 191 

Burcq,  effect  of  metallic  copper  on  bacteria. . .  250 
Biitschli,  nature  of  central  body  in  bacteria.   216, 217 

Butter,  bacteria  in 259 

Cabbage,  natural  infection  in. . .  92, 93, 94,  102, 124 

Cages  for  inoculation 108 

Calcium  carbonate,  effect  of,  on  longevity 61 

Calcium  chloride,  effect  on  luminous  bacteria.     60 
Camera, 

Abbe,  for  drawing 130 

centering  of  horizontal 137 

enlarging,  reducing,  and  copying 146 

photographing  with  horizontal 137 

Cankers  produced  by  bacteria 8 

Capillary  drop  flask 21, 226 

Capsule, 
branching  caused  by  one-sided  development 

Of 221 

examination  of  unstained 19 

Friedlaender's  stain 220 

methods  of  staining  for  demonstration  of .   19, 194 

Moore's  contrast  stain 221 

Carbol-fuchsin  187 

Carbol-methylene  blue 188 

Carbolic  acid 74 

for  sterilizing  surface  of  diseased  material . .     14 
Carbon  dioxide, 

assimilation  of,  by  bacteria 64 

behavior  in 59, 223 

cultures  in,  factors  to  be  considered 59 

generation  of 54, 55 


Page. 
Carbon  dioxide — continued. 

removal  of  all  oxygen  from 57 

test  for,  in   fermentation-tubes 61 

test  for  purity  of 55 

washing  of 55 

Carbon  monoxide,  effect  on  bacteria 58 

Carney's  fixing  fluid 202 

Casein, 

non-precipitated,  clearing  of 267 

precipitated,  re-solution  of 46 

separation  of,  in  milk-cultures 46, 67 

Cavities  produced  by  bacteria 8 

Cell-nucleus   216,  217, 219 

Cellulose, 

in  bacteria 219,  254 

pure,  fermented  by  bacteria 209 

Chains,  occurrence  of  bacteria  In 22 

Chamberland  autoclave 85 

Chamberland  filter, 

cleaning  of 45, 227 

enzymes  entangled  in  walls  of 68 

penetration  of,  by  bacteria 45, 226 

used  in  study  of  antagonism 73 

Characters, 

cultural,  value  of 178 

decimal  system  of  recording 175 

morphological,  value  of 176 

variation  in,  due  to  environment 182, 183 

Check-plants, 

importance  of 14,  186 

proper  behavior  of 15,  16 

Checking  work,  necessity  for,  and  methods  of.  184 

Chemicals,  for  use  in  culture-media 97 

Chemotropism  27 

Chester, 

decimal  system 175 

terminology  for  description  of  colonies 30 

Chlamydothrix  162 

Chloroform 74,  250,  254 

Chlorozinc-iodide, 

use  in  staining  anthrax  spores 218 

Chodat,  lactic  acid  bacteria  in  cheese 260 

Cholera  vibrio, 

cultivation  on  potato 249 

destroyed  by  river  waters  in  India 251 

effect  of  copper  on 250 

method  of  diagnosing  and  isolating 229 

names  for 173 

Chondromyces  165 

Chromatium  164 

Chromatium  Okenii,  cells  connected  sidewise.  216 
Chromic  acid, 

cleaning  mixture 200 

use  in  staining  spores 218 

Chromo-aceto-osmic  acid  for  fixing  fluid 202 

"  Chromo-agars  "  for  differentiation 229 

Chromogens,  medium  for  study  of 48, 224 

Chromoparous  bacteria 237 

Cladothrix 162 

Classifications  154 

Classification, 

iMigula's  system 159 

Fischer's  system 157 


INDEX. 


273 


Page. 
Classification — continued. 

water-bacteria  203 

Winslow  &  Rogers 265 

with  reference  to  nitrogen-nutrition 175 

Cleaning,  glassware 100,  226,  227 

Cleaning  mixture,  chromic  acid 200 

Clostridiutn  pasteurianum,  separation  by  heat.  106 

Cloth,  impervious  to  light 143 

Clouding, 

nature  of,  in  culture-fluids 42 

rapidity  of,  in  culture-fluids 42 

Coccaceae,  revision  of 265 

Coccobacteria  septica 176 

Coconut-water,  as  a  culture-medium 41 

Cocothrix  172 

Cohn, 

Bacillus  of 158 

Bacterium  of 166 

nutrient  solution  of 197 

rise  of  temperature  in  cotton-wool  waste. . .  248 

Collections,  the  making  of 117 

Collodion, 

sac-method  for  study  of  antagonism 73 

sacks,  preparation  of,  for  dialyzing 37, 229 

Colonies, 
acid-forming,  special  agar  for  detection  of.     51 

characters  to  be  noted  on  solid  media 29 

counting  of 36 

effect  of  physical  conditions  on  appearance 

of 182, 183 

measurement  of 115 

photographing  of 134 

Color-scales, 

importance  of 109 

kinds  recommended 263 

Comma  bacillus, 

classification  of 173 

Koch's  paper  on 212 

Conditions   which  are  unfavorable,   influence 

on  cell-unions 22 

Conrad,  fermentation  of  sauerkraut 257 

Constancy  of  characters 183 

Contact-irritation   27 

Contamination,  how  to  avoid 103 

Control-plants,  proper  behavior  of 15,  16 

Coplin's  staining  jar 119 

Copper-chrom-filter  201 

Copper  salts,  germicidal  value  of 251 

Copper  sulphate 74 

Corallin,  test  for  slime  derived  from  starch..  221 
Cotton, 

dry  heating  in  bulk 99, 101 

natural  infection  in 95 

surgeons'  absorbent,  sterilization  of 101 

waste,  for  laboratory  use 107 

Counting-plates,  ruled 36, 227 

Cover-glasses,  cleaning  of 227 

Cover-glass  preparations 20, 28 

Covers,  discarded,  care  of 107 

Cramer,    resistance    of    spores    to    dry    heat, 

cause  of 218 

Crenothrix 162 

Cristiani,  bacteria  in  upper  air 255 


Page. 

Cross-inoculations  186 

Cross-level  149 

Crystals, 

formation  of,  in  milk-cultures 46 

in  old  cultures 66 

prevent  making  thin  sections 122 

Cucumber,  natural  infection  in 92,178,215 

Cultural  characters, 

value  of 178 

variations  in,  due  to  environment 182, 183 

Culture-fluids, 

cold  sterilization  of 43,  52 

examination  of,  before  inoculation 42 

growth  in,  characters  of,  to  be  noted 42 

non-synthetic 195 

synthetic 197 

time  required  for  clouding 42 

Culture-media  223 

agar-nutrient 31,  195 

beef-broth 45, 195 

blood-serum  48 

containing  sugars,  sterilization  of 98 

cooked  vegetables  for 40 

Dunham's  solution 195 

egg-albumen 48 

egg-yolk 49 

for  luminous  bacteria 60, 198 

gelatin  29,  196 

hydrochloric  acid  in 98 

litmus  milk 48, 196 

milk 46 

non-synthetic  195 

plant  juices  used  as 41 

preparation  and  care  of 97 

protection  from  light 53 

raw,  preparation  of 41 

record-book  for 109 

silicate-jelly 36,  198 

special,  for  differential  purposes 229 

starch-jelly  50, 196 

sterilization  of,  in  autoclave 98 

storage  of 99 

synthetic  197 

titration  of 69 

Culture-rooms  104 

Cultures, 

elective 106 

involution  forms  in 23 

method  of  keeping  notes  on 113 

methods  of  testing  purity 184, 185 

plate,  stab  and  streak 29, 30 

preservation  of,  in  museums 262 

storage  of  stock 123, 263 

treatment  of  spilled 107 

Cyanophyceae,  cilia  on  side  walls  of 211 

Cystobacter 165, 218 

Cytase  67 

Dallinger,  measurement  of  unstained  flagella, 

170, 219 

Dallinger  &  Drysdale,  Bacterium  termo,  fla- 
gella of 20 

Dangeard,  green  bacteria 237 

Dannappel,  spores  sensitive  to  heat 22, 246 


274 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Page. 

Dark-room, 

arrangement  of 15° 

ventilation  of 149 

d'Arsonval, 

negative  results  with  ozone 252 

osmotic  pressure  of  bacteria 247 

Darwin's  wax-mixture 200 

Dawson,  root-tubercles  of  legumes 64 

Davenport  &   Castle,   acclimatization   to  high 

temperatures  222 

Decimal  system  of  recording  characters 175 

de  Freudenreich, 

bacteria  in  mountain  air 255 

penetration  of  Chamberland   filter  by  bac- 
teria     226 

Degeneration-forms  23, 177, 252 

de  Koninck,  gas-generator 54 

de  Lagerheim,  macaroni  as  a  culture-medium.  224 
Delbrueck,  acid  fruits,  effect  on  cholera  or- 
ganism    249 

Delphinium,  bacterial  disease  of 92,  139 

Denitrifying  organisms,   Giltay   &   Aberson's 

culture-medium  for 198 

Desiccation,  effect  of 70, 248 

Developer, 

hydrochinon 146,  201 

lantern-slide  201 

ortol 140 

pyrogallol  200, 201 

Development  of  negatives 140 

Dewar  glasses,  for  liquid  air 80 

Dextrin  for  use  in  fermentation-tubes 52 

Dialyzing,  collodion  sacks  for 37 

Diastase 67 

Diastasic  action,  medium  for  study  of 50,  196 

Dieudonne,  part  of  spectrum  harmful  to  bac- 
teria    244 

Differentiation  of  species 25,225,229,230 

Dilution  method  for  obtaining  pure  cultures.  .  226 

Diphenylamine-test  for  nitrates 267 

Direct-infection  experiments,  value  of 9 

Disease, 

conditions  favoring  the  spread  of 93 

geographical  distribution  of 7 

meaning  of  term 4 

signs  of 7 

varietal  resistance  to 93, 186 


Disinfectants 


74 


for  sterilizing  surface  of  diseased  material. .     14 


literature  of 


250 


Disinfection  of  hands,  wounds,  floors,  tables, 

etc 107 

Distilled  water, 

of  a  high  degree  of  purity 129 

made  in  large  quantities 124 

made  in  small  quantities 128 

storage  of I26, 129 

Distribution,  geographical   7 

Divided-plate   method   for   study   of   antago- 
nism    73 

Division  of  cells,  arrangement  after 22,  218 

Downes  &  Blunt,  discover  germicidal  action 

of  light 243 


Page. 

Drawings,  on  photographic  prints 151 

Dreyer,  staining  of  bacteria  in  tissues 222 

Dreyfuss,  cellulose  in  bacteria 254 

Dry  air,  sensitiveness  of  bacteria  to 71,249 

Dubois,  fluid  medium  for  luminous  bacteria. .  242 

Duckwall,  method  of  staining  flagella 193 

Dunham's  solution 19=; 

Eau  de  Javelle 207 

Economic  aspects, 

conditions  favoring  spread  of  disease 91 

methods  of  prevention 93 

natural  methods  of  infection 91 

need  of  statistics  relating  to  plant  diseases . .     90 
Egg-albumen, 

for  culture-media 48 

Mayer's  i  ig 

Eggs  as  a  culture-medium 224,  225 

Egg-yolk,  for  culture-medium 49 

Ehrenberg, 

Bacterium  of 165-169 

Spirillum  of 173 

Ehrlich's  anilin-water  gentian  violet 187 

Ehrlich-Weigert  anilin  methyl  violet 187,  188 

Elective  culture 106 

Embedding,    method    of,    for    microtome-sec- 
tions      1 18 

Engelmann, 

green  bacteria 236 

photometric  bacteria 243 

Enlargements,  photographic 147 

Environment, 

influence  on  cultural  characters .    178,  182,  183, 222 
response  morphologically  to  change  in .   176,  222 
Enzymes, 

filtering  of 68 

kinds  of 66 

nature  of 66 

not  in j  ured  by  dry  heat 68 

number  produced  by  one  organism 68 

substances  inhibiting  action  of 68 

thermal  relations  of 67, 233 

Errera, 

India  ink  in  microscopy 226 

spirillum  of  large  size 19 

Error,  methods  of  guarding  against 184 

Ether,  sulphuric 74,  254 

Euspirosoma  161 

Ewart,  green  bacteria 238 

Ewell,  anaerobic  apparatus 56 

Exposure  scales 143 

Exposure,  time  of, 

for  isochromatic  plates 143 

for  lantern  slide 146 

for  Petri-dish  poured  plates 134 

for  photomicrographs 143 

for  velox  paper 151 

Faber's  pencils  for  glass no 

Farmer's  reducing  solution 142 

Favorable  influence  of  one  organism  on  an- 
other     72,74 

Fehling's  solution, 

reaction  of  peptone  with 229 

when  not  usable 208,  234 


INDEX. 


275 


Pane. 

Feinberg,  on  existence  of  nucleus 217 

Fermentation, 

causes  of 60 

commercial  importance  of 60 

determination  of  gases  produced  by 61 

effect  of  calcium  carbonate  on 6r 

isolation  of  products  of 61 

literature  of 232 

meaning  of  term 60 

measurement  of  volume  of  gas  produced.  . .     61 

observations  to  be  made  on 61 

of  cellulose 209 

pectic  257 

products  of 60,  or 

Fermentation-tubes, 

absorption  of  air  into  closed  end  of 52 

fluids  to  be  used  in 52 

observations  to  be  made  on  cultures  in 52 

style  preferred 53 

substances  to  be  tested  in 52 

test  for  presence  of  air  in  closed  end 54 

wooden  rack  for 52 

Fermented  foods,  bacteria  of 235 

Fermi, 
concentrated  solution  for  silicate-jelly...  39,  197 

effect  of  acids,  etc.,  on  bacteria 249 

studies  of  ferment-bacteria 233 

Fermi  und  Montesano,  reagent  for  sugar 234 

Fermi  und  Pernossi,  action  of  heat,  light,  etc., 

on  enzymes 233 

Ferran,  aerobic  behavior  of  tetanus 231 

Picker,    glass,    influence    of    substances    dis- 
solved out  of 223 

Filaments,  bacteria  in  form  of 22 

Filters, 

Berkefeld  and  Chamberland,  for  cold  steril- 
ization of  fluids 43 

Zettnow's  copper-chrom 201 

Filter-papers, 

enzymes  held  in  meshes  of 68 

most  convenient  form  of 34 

reaction  of 34 

Filtration, 

by  means  of  compressed  air,  etc 44 

inside-out  method 43,  226 

of  milk  for  culture-media 46 

of  nutrient-agar 33-35 

of  thick  fluids 69 

Fiocca's  spore-stain 194 

Fischer,  A., 

classification  of 157 

experiments  on  plasmolysis 22, 254 

flagella-stain  191 

subfamilies  distinguished  by  flagella 20 

Fischer,  B., 

bacteria  in  sea-air 255 

germicidal  action  of  light  through  water 244 

Fish-parasites  213,214 

Fixation  of  plant  material  containing  bacteria.      8 

Fixing  fluids 202 

Flagella  219 

classifications  based  on 20,  157 

earliest  demonstrations  of 20 

stains  20,  190 


Page. 

Flagel  la-staining, 

cultures  best  suited  for 190 

literature  on 219 

technique  189 

Flasks,  for  storing  distilled  water 120 

Flax,  retting  of 257, 258 

Flemming, 

fixing  fluid 202 

triple  stain 188 

Flexner's  anilin  gentian  violet 187 

Flocculence  in  fluid-cultures 22 

Florideae,  source  of  agar 32,  224 

Fluorescine  65 

Focus-difference,  with  achromatic  objectives.  140 
Foerster,  Chromatium  Okenii,  cells  connected 

sidewise  • 216 

Folmer  &  Schwing,  enlarging,  reducing,  and 

copying  camera 146 

Formaldehyd   74 

Formalin   74 

differential  effect  on  bacteria 229 

germicidal  action 252, 253 

preservation  of  cultures  by  use  of 263 

Formulae  187 

Forster,  gelatin  with  high  melting  point 225 

Foth's  spore-stain 218 

Fractional  method  of  culture. 226 

Fraenkel  &  Voges'  solution 197 

Frankland, 

effect  of  gases  on  bacteria 231 

photographic  action  of  bacteria 242 

Frankland  &  Ward,  method  for  study  of  an- 
tagonism      73 

Freer  &  Novy,  germicidal  action  of  organic 

peroxides  253 

Freezing 246 

effect  on  anthrax,  etc 247 

effect  on  B.  typhosus 247, 258 

effect  on  bacteria  of  pest  and  diphtheria 247 

osmotic  pressure  a  factor  in 247 

with  liquid  air 79-83 

with  salt  and  ice 83 

Frost, 

antagonism  73 

gasometer  61 

Friedlaender, 

acid-fast  stain 187 

capsule-stain  220 

Fribes,  retting  of  flax 257 

Fruit-ether-forming  bacteria 259 

Fuchsin,  anilin 187 

Fuller's  scale 69 

Funck,  cleaning  of  cover-glasses 227 

Fiirbringer     und     Freyhan,     disinfection     of 

hands 252 

Gabbett's  stain 188 

Gas-analysis  55 

Gases,  compressed 81 

Gas-generator, 

de  Koninck 54 

hydrogen,  with  wash-bottles 55 

Kipp 54 

Gasometer,  Frost  s 61 

Gas-pressure  regulator,  Murrill's 78 


276 


BACTERIA    IN    RELATION   TO    PLANT   DISEASES. 


Page. 

Gauze,  surgeon's,  uses  in  laboratory 102 

Gelatin, 

antiseptic  salts  in 3° 

brands     recommended     for    bacteriological 

work 3 r 

composition  of 30 

impurities  in 30 

melting  point  of 30 

nitrates  in 224 

per  cent  recommended  for  media 30 

plate-cultures,  stabs,  streaks 29 

standard  nutrient,  preparation  of 196 

sterilization  of 98 

variation  of  same  organism  on 180 

with  cane-sugar 51 

with  high  melting  point 225 

with  malic  acid 51 

with  soluble  starch,  etc 51 

Gelatinization,  of  old  milk-cultures 46 

Gelidium,  species,  source  of  agar-agar 31 

Generic  characters  based  on  morphology 156 

Generic  names  rejected 174 

Gentian  violet, 

Ehrlich's  anilin-water 187 

Flexner's  anilin 187 

Gerlach,  germicidal  action  of  lysol 251 

Germicides 74 

literature  on 250 

Germination,  spores  should  be  subjected  to. . .     21 

Gessard,  green-fluorescent  bacteria 237 

Giltay  &  Aberson,  culture-medium  for  denitri- 
fying organisms 198 

Glassware, 

cleaning  and  sterilization  of 100 

pencils  for  writing  on in 

solubility  of 129, 223 

Globig,  thermophilic  bacteria 248 

Glue,  blue-pigment  bacteria  injurious  to 237 

Glycerin, 

agar  33, 196 

gelatin  262 

mounts,  sealing  of 262 

Goriansky,  wood-vinegar  as  a  disinfectant. . . .  252 

Gram's  stain 188 

Nicolle's  modification • 222 

Grape-sugar 

harmful  effect  of 223 

test  for,  in  plant  tissues 208 

Green,  germicidal  value  of  copper  salts 251 

Green  bacteria 237, 238 

Green-fluorescent  bacteria 65 

Gessard's  studies 237 

Jordan's  studies 238 

Thumm's  studies 238 

Griess-Ilosvay's  reagent 229 

Grimbert,  on  value  of  synthetic  media 51 

Growth  of  bacteria, 

manner  and  rapidity  of 27 

on  media  exhausted  for  other  organisms. . .  211 

rates  of,  in  fluid  cultures 42 

restrained  by  acid  gelatin 30 

Grove,  glycerin  mounts  rendered  air-tight 262 

Gruber,  resistance  of  spores 246 


Page. 
Guillebeau    &    de    Freudenreich,    agar    made 

without  filtering 223 

Haegler,  oentrifuging  agar 225 

Halation,  means  of  avoiding 136 

Hallier,  views  concerning  morphology 176 

Hands,  disinfection  of 252 

Hanging-drop  cultures 22 

containing  single  organism 27 

manner   and   rapidity   of   growth   at  given 

temperatures  27 

Hankin,  germicidal  action  of  river  water 251 

Harris,  collodion  sacs,  method  of  making 229 

Haswell,  method  of  substituting  alcohol   for 

water  226 

Hauser, 

spore-stain  194 

preservation  of  cultures  for  museums 263 

Heat, 

as  a  means  of  separating  organisms 106 

as  a  sterilizer  for  surface  of  tissues 13 

dry,  cause  of  resistance  of  spores  to 218 

effect  on  pigmentation,  pathogenicity,  spor- 

ulation  87 

enzymes  injured  by  moist 67 

influence  of  substratum  on  resistance  to.  219,  247 

Heidenhain's  iron-haematoxylin 189 

Heim,  anaerobic  cultures  of 231 

Hellstroem,  grape-sugar,  harmful  effect  of .  . .  223 
Hempel, 

burette  for  gas-analysis 55 

pipette  for  liquid  reagents 55 

Henle,  on  proofs  of  pathogenesis 9 

Herter  &  Foster's  test  for  indol 201 

Hesse, 

media  for  water-bacteria 196,  258 

method  of  cultivating  anaerobes 230 

Hill's  hanging-block  method 22,  228 

Hinterberger,  method  of  staining  flagella 220 

Hiss,  media  of, 

for  differentiation  of  motile  bacteria 26 

for   differentiation   of   typhoid,   colon,   and 

allied  bacilli 230 

Kitchens,  on  autoclaving  bouillon  containing 

sugar  99 

Hoffmann,  infiltration  in  vacuo 226 

Hoods, 

improvised  104 

in  laboratories 104 

Host,  color  changes  in, 
due  to   injury  other  than   that  caused   by 

parasite  66 

due  to  oxidation  phenomena 65 

due  to  parasite 65 

Host,  reaction  to  parasitic  attack 8 

Host-plants,  kinds  attacked  by  a  single  para- 
site     87 

Hydrochloric  acid,  in  culture-media 98 

Hydrogen, 

compressed,  where  obtained 81 

cultures  in 56,  57 

test  for,  in  fermentation-tubes 61 

Hydrogen-cellulose  ferment 106, 209 


INDEX. 


Page. 

Hydrogen  generated  with  zinc 54,  SS 

hydrogen    sulphide   in,  means    of  avoiding 

evolution  of 56 

impurities  in 55 

removal  of  last  traces  of  oxygen  from 57 

washing  of 56,  57 

Hydrogen  peroxide, 

use  in  staining  spores 218 

substance     causing     liberation     of     oxygen 

from  67, 234 

Hydrogen  sulphide 62,  242,  243 

Hypo  for  fixing 142 

Tee,  bacteria  in 247,  258 

India  oil-stone 121 

Indigo-carmine,  reduction  of,  by  bacteria 62 

Indigo-blue,  production  of 257 

Indol, 

production  of,  media  suitable  for 62 

tests  for 62, 201 

Infection, 

carriers  of 91, 178,  215 

conditions  favoring 16 

due  to  ultramicroscopical  organisms....   18,211 

natural  methods  of 8, 92 

Infection,  stomatal 84,  86,  90,  92,  108,  126 

Infection-experiments,  value  of  direct 9 

Infectious  material,  final  disposal  of 106 

Infiltration, 

in  vacuo 8, 226 

with  paraffin 118 

Injection-needle 101, 227 

Inoculated  organism,  method  of  proving  viru- 
lence     16,  185 

Inoculated  plants,  labeling  of 112 

Inoculation, 

cages 108 

checks  on 16,  186 

cross  186 

methods  of 108 

with  more  or  less  exact  numbers  of  bac- 
teria    226 

Inoculation-experiments, 

conditions  favoring 16 

how  carried  on 15 

importance  of 14 

where  best  made 16 

with  mixed  organisms 9,  72 

Insects,  inoculation  by  means  of. .  91,  108, 178,215 

Instruments,  sterilization  of 100, 107 

Intensifying  negatives 142 

Invertase  66 

Investigation,  training  necessary  for 181 

Involution-forms  23,  177 

Iodine-starch  test  for  nitrites 63 

Iron-haematoxylin 188,  189 

Irritation,  contact 27 

Isochromatic  plates, 

for  photographing  stained  sections 140 

time  of  exposure  with 143 

uses  of 136 

Isolation  of  bacteria, 

from  diseased  tissues,  methods  of n 

on  slant  agar,  etc 227 


Page. 

Twanoff,  penetrating  power  of  formalin 252 

Jeffer,  ruled  counting  plate 36 

Jennings,  contact-irritation 27,  254 

Johan-Olsen,  branching  forms 217 

Jolles   und   Winkler,   bacteria   in   oleomarga- 
rine and  butter 259 

Jones,  enzymes  from  soft-rot  organisms 68 

Jordan,  pigments  produced  by  green-fluores- 
cent bacteria 65 

Kabrhel,  methylen  blue  as  oxygen  indicator. .  232 

Kaiser,  glycerin-gelatin 262 

Karten,  chemical  analysis  of  agar-agar 32 

Kasansky,  effect  of  winter  cold  on  bacteria.  . .  247 

Kaufmann,  method  of  staining  capsules 194 

Kedrowski,  anaerobes  grown  with  aerobes. . . .  231 

Kendall,  decimal  system 176 

Kephir 214 

Kern,  two  endospores  in  a  cell 22 

Kipp  gas-generator 54 

Kitasato  flask 68 

Klebs,  fractional  method  of  culture 226 

Klein,  A., 

anaerobic  apparatus  of 232 

spore-stain  219 

Klein,  L.,  bacilli  bearing  green  spores 218 

Klepzoff,  freezing  experiments  of 246 

Knauer,  cleaning  slides  and  cover-glasses....  226 

Knives,  microtome 122 

Koch,  A.,  branching  of  B.  pediculatum 221 

Koch,  R., 

comma  bacillus  of 173 

demonstration  of  flagella  by  stains 20 

importance  of  photography  in  microscopic 

works   23 

magnification  recommended  for  photomicro- 
graphs       24 

methods  of  isolating  parasite  from  tissue,  n,  226 

paper  on  tuberculosis 153 

rules  of  proof 18, 226 

streak-method  of  isolating  bacteria u,  226 

views  concerning  morphology 176 

Konradi,  germicidal  action  of  soaps 253 

Kraemer,  germicidal  action  of  copper 253 

Kratschmer  und  Niemelowicz,  stringy  bread . .  260 

Krause,  Bacillus  pyocyaneus 223 

Krueckmann,    preservation    of    cultures    for 

museums  263 

Kuehne, 

carbol  methylene  blue 188, 222 

malachite  green 222 

Ktintsler,  animal  origin  of  bacteria 264 

Kuntze,  medium  for  Bacillus  prodigiosus 238 

Kuprianow,  device  for  filling  test  tubes 227 

Labels,  for  tubes,  flasks,  etc no,  in 

Lab  ferment 67 

Laboratory, 

care  of 96 

equipment  of 94 

things  to  be  considered  in  location  of 94 

Lamella,  middle,  dissolution  of 67 

Lamprocystis   163 

Lantern-slides, 

apparatus  for  making 143,  146 


278 


BACTERIA    IN    RELATION    TO    PLANT    DISEASES. 


Page. 

Lantern-slides — continued. 

development  of 146,  201 

exposure  of 146 

Larkspur,  natural  infection  in 92,  139 

Laser,  on  action  of  hydrogen  peroxide 234 

Lautenschlager  oven 100 

Lead  acetate  paper 62 

Leguminosae, 
root-tubercle  bacilli  of,  fixation  of  nitrogen 

by  64 

Moore's  culture-medium  for 197 

Lenses, 

achromatic,  defects  of 14° 

apochromatic,  in  photomicrography 139 

for  bacteriological  work 23 

Lenses,  photographic, 

Goerz  130 

Voigtlaender  130 

Zeiss  Double- Protar 130, 132 

Zeiss  Planar 132 

Zeiss  Protar 132 

Zeiss  Unar 132 

Lenticels,  natural  infection  through 92 

Lepeschkin,  branched  form  described 218 

Leucin  63 

Leuconostoc  mesenterioides,  isolation  of 106 

Levin,  bacteria  rare  in  Arctic 255; 

Levy,  physical  properties  of  enzymes 68 

Lewandowski,  test  for  phenol 63 

Liborius,  oxygen  in  depths  of  agar 230 

Light, 

action  on  bacteria 71, 243,  244 

bactericidal    action    ascribed    to    hydrogen 

peroxide  244 

germicidal    action   on    dust   greatest   when 

moistened   244 

germicidal   in  hydrogen 244 

germicidal  action  in  water 244 

Light  filter  for  photomicrography, 

dry 24 

Zettnow's   24, 201,  263 

Lilac  blight 64 

Liquefaction  in  gelatin  stab-cultures, 

characters  to  be  noted 29 

interferences  of  various  substances  with.  ...     29 

Liquid  air,  effect  of,  on  bacteria 79, 247 

temperature  of 80 

Lister,    dilution   method    for   obtaining   pure 

cultures  27, 226 

Litmus-lactose-agar  33,  196 

Litmus  milk, 

cultures  in,  observations  on 48 

preparation  of 48, 196 

Litmus  paper,  for  testing  acidity 61 

Litmus,  reduction  of,  by  bacteria 62 

Loeb,  typhoid  bacillus,  branching  forms  in...  218 
Loeffler, 

alkaline  methylene  blue 188 

extraction  of  enzymes 68 

flagella-stain  icjo 

Loew, 

culture-medium  for  Bacillus  prodigiosus. . .  239 
enzymic  fermentation  of  tobacco 257 


Page. 

London,  resistance  to  starvation 223 

Loops,  platinum-indium,  for  making  transfers .      43 

Loquat,  bacterial  disease  of 88 

L6 wit's  flagella-stain 192 

Luminous  bacteria 60,  241 

cause  of  luminosity  of 60 

culture-medium  for 198 

Dubois'  fluid  medium  for 242 

effect  of  chlorides  on 60 

effect  of  magnesium  sulfate  on 60 

effect  of  manganese  sulfate  on 60 

effect  of  potassium  iodide  on 60 

effect  of  potassium  nitrate  on 60 

effect  of  potassium  sulfate  on 60 

in  or  near  salt  water 60 

in  sand-fleas 60 

Molisch's  work  on 60 

on  meat  exposed  in  markets 60 

on  synthetic  media 60 

spectrum  of 242 

thermal  relations  of 60 

Lunt,    stock-cultures   of   water-bacteria,    how 

best  kept 263 

Lysol, 

for  sterilizing  surface  of  diseased  material.     14 

germicidal  action  of 251 

Maassen, 

branching  forms  produced  at  will  by 23 

culture-fluid  of 198 

fruit-ether-forming  bacteria 259 

reducing  powers  of  bacteria 62,  243 

Mace, 
recuperation  of  pigmentation  and  pathogen- 

icity 237 

ruled  counting-plate 36 

Macfadyen, 

effect  of  liquid  air  on  bacteria 247 

thermophilic  bacteria 248 

Mackensie's  culture-fluid 198 

Magnesium  chloride,  effect  on  luminous  bac- 
teria       60 

Magnesium  sulfate,  effect  on  luminous  bac- 
teria       60 

Magnification,  determination  of 115 

Maize, 

artificial  infection  of 90 

cross-section  of  stem 4 

natural  infection  in 92 

Malachite  green,  use  of 189, 222 

Maltose-agar  33 

Manganese   sulfate,   effect  on   luminous  bac- 
teria        60 

Marmier   et   Abraham,   germicidal   action   of 

ozone  252 

Marpmann, 

"  chromo-agars  "  for  differentiating 229 

germicidal  action  of  sodium  fluoride 253 

iron  bacteria 261 

Marsh  gas 61,209 

Maximum  temperature, 

how  determined 75 

range  of 87 

Mayer's  culture-fluid 197 

Maze,  root-tubercle  bacteria 240 


INDEX. 


279 


Page. 

Meat-extracts,  bacteria  in 260 

Media  for  cultures, 

agar 31, 195 

animal  fluids 45 

gelatin 29,  196 

milk 46, 196 

non-synthetic  195 

plant  juices 41 

silicate-jelly  41 

special  49 

starch-jelly  5°.  T96 

sterilization  of 85 

synthetic    49,  197 

vegetable  media,  solid 39 

Media  free  from  spores,  easy  sterilization  of.  40,85 

Mercaptan   62, 243 

Mercuric  chloride 74 

for  sterilizing  plants  before  inoculation.  108,  109 
for  sterilizing  surface  of  diseased  material.     14 

germicidal  action  of 232 

in  hot  alcohol,  as  a  fixing  fluid 8 

preservation  of  cultures  by  use  of 263 

Messea,  classification  of 20, 264 

Mctcalf,  bacillus  softening  agar 32 

Meters,  for  photographic  exposures 143 

Methane-cellulose  ferment 106 

Methods  of  work, 

checking 184 

literature  on 226 

Methylene  blue, 

as  a  test  for  free  oxygen 57, 232 

Loeffler's  alkaline 188 

reduction  of,  by  bacteria 62,  239 

Methyl  violet,  as  a  germicide 74 

Meyer,  effect  of  liquid  air  on  bacteria 247 

Michaelis,  thermophilic  bacteria 248 

Micrococcus  160 

Micrococcus  progrediens,  small  size  of 18 

Micrometers,  stage 115 

Micro-organisms,  cell-life  without 211 

Microscope, 

achromatic  objectives  for 140, 265 

apochromatic  objectives  for 130 

for   recording   exact   location   of   desirable 

fields  129 

Leitz 129 

oculars  130 

Spencer  129 

Zeiss,  for  bacteriological  investigations 129 

Zeiss,  for  photomicrographic  work 129 

Microspira 161, 172 

Microtome,  kind  preferred 122 

Microtome-knives 122 

Microtome-sections, 

cutting  and  care  of 119 

mounting  of 119 

staining  of 119, 120 

Microzymas  of  Bechamp 176 

Migula,  classification  of 159 

Milk, 

a  good  culture-medium 47 

anaerobes  in 46 


Page. 
Milk — continued. 

clearing  of 46, 267 

coagulation  of,  by  bacteria 67 

enormous  numbers  of  bacteria  in 196 

litmus  196 

observations  to  be  made  on  cultures  in 46 

preparation  of,  for  culture-medium 46 

reddened  by  bacteria 259 

resistant  spores  in 46 

ropiness  in 259,  260 

sterilization  of 46, 98 

temperature    governs    bacterial    multiplica- 
tion in 260 

Minervini,  bactericidal  action  of  alcohol 252 

Minimum  temperature, 

for  growth,  how  determined 76 

range  of 87 

Miquel, 

bacteria  in  sea-air 255 

thermophilic  bacteria 247, 248 

Mixed  cultures,  behavior  of 72 

Moller's  spore-stain 218 

Molisch,  on  luminous  bacteria 60 

Monas 173 

Moore,  A.,  capsule-stain 221 

Moore,  G.  T., 

copper  sulphate  as  a  germicide 74 

culture-medium  for  root-tubercle  bacilli.  ...   197 

root-tubercle  bacteria 241 

soil-inoculation  for  legumes 64 

Moore,  V.  A., 

flagella-stain  190 

Moore  &  Kellerman,  action  of  copper  on  B. 

typhosus  in  water 97 

Morphological  characters, 

change  in,  due  to  changed  environment 222 

value  of 176 

Morphology  19 

extreme  views  concerning 176 

general  account  of 18 

generic  characters  should  be  based  on 156 

insufficient  for  differentiation  of  many  spe- 
cies       25 

literature  on 215 

modern  views  concerning 177 

Morton,  flagella-stain 220 

Motility  of  bacteria 26 

Motor-reflex  in  bacteria 254 

Mueller,  bacterial  reduction  processes 239 

Miiller,  genus  Vibrio 172 

Muir,  capsule-stain 194 

Murrill,  gas-pressure  regulator 78 

Museums,  preservation  of  cultures  for. . .  262,  263 

Mustard,  natural  infection  in 92 

Mycobacterium   172 

Myconostoc  161 

Myxobacter  165 

Myxobacteriaceae,  characters  of 164 

Myxococcus 165 

Naegeli's  nutrient  solution 197 

Nakanishi,  nucleus,  existence  of,  in  bacteria. .  217 

Nectaries,  natural  infection  through 92 

Needle-punctures,  inoculation  by 108 


280 


BACTERIA   IN    RELATION   TO    PLANT   DISEASES. 


Page. 

Needles,  platinum-iridium,  for  making  trans- 
fers       43 

Negatives, 

development  of 14° 

records  on 142 

Nessler*s  test,  how  used 61 

Neisser's  spore-stain 104 

Neumann,  variability  of  pigment- formation . . .  238 

Neutral  red 230 

Nicolle,  modification  of  Gram's  method 222 

Night-blue, 

capsule-stain 221 

stain  for  flagella 220 

Nitrate  bacteria, 

fluid  culture-medium  for  isolation  of 199 

nutrient  agar  for  isolation  of 109 

Nitrate  bouillon 63 

diphenylamin  test 267 

Nitrates,  reduction  of,  test  for 63 

Nitrifying  organisms,  magnesia-gypsum  blocks 

for 200 

Nitrite  bacteria,  fluid  medium  for  isolation  of.  199 

Nitrites,  test  for 63 

Nitro-bacteria  175 

Nitrogen-assimilating    soil-bacteria,    medium 

for  199 

Nitrogen-bacteria  175 

Nitrogen-free  media 51, 198 

Nitrogen, 

fixation  of,  by  bacteria 64 

growth  in,  apparatus  for  testing 58 

removal  of  oxygen  from 57 

Nitrogen-nutrition,  a  basis  for  classification..  175 

Nitrogen  salts,  in  special  media 51, 197 

Nitromonas  240 

Nitrous  and  nitrate  bacteria 175 

Nomenclature  154 

Non-halation  plates,  uses  of 136 

Novy, 

a  new  thermoregulator 228 

jar  for  anaerobic  cultures 56,  57, 58, 231 

Nucleus,  existence  of,  in  bacteria.   159,216,217,219 
Nutrient   material,    amount   needed   by   bac- 
teria     225 

Objectives,  achromatic, 

disadvantages  of 140 

of  excellent  definition 265 

Objectives,  apochromatic, 

Spencer  16  mm 140 

Zeiss  130 

Occlusion  of  vascular  system  due  to  bacteria . .    8, 12 

Oculars  130 

for  photomicrography 139 

Ohlmuller,  germicidal  action  of  ozone ^51 

Oleomargarine,  bacteria  in 259 

Olive-knot  organism, 

artificial  inoculations 10 

crystals  produced  by 66 

optimum  temperature  for 85 

Omelianski, 

blocks  for  nitrifying  organisms 200 

isolation  of  hydrogen-cellulose  ferment.  106, 209 

sodium-formate  medium 50 

spore  readily  stained  by  anilin  dyes 22 


Page. 
Optimum   reaction   of  medium   for  bacterial 

growth  69 

Optimum  temperatures 75,  85 

Origin  of  bacteria 177 

Ortol  developer 140 

Ostwald-Pfeffer  water-bath 78 

Oven,  paraffin no 

Oversteaming,  effect  of 98 

Oxidases   67 

Oxidation,  pigments  due  to 66 

Oxygen, 

compressed,  where  obtained 81 

little,  in  deep  layers  of  agar 230 

pigment- formation,  dependent  on 238.  230 

removal  from  hydrogen,  nitrogen,  and  car- 
bon dioxide 57 

Oxygen,  relation  of  bacteria  to,  in, 

fermentation-tubes  52 

surface  and  deep  growths 51 

Ozone,  action  on  bacteria 251,  252,  253 

Pacinia 172 

Pake,  ruled  counting-plate 36 

Paper, 

blue-print,  for  pen-and-ink  work 150 

drawing-board  266 

salted  silver,  for  pen-and-ink  work 150 

Paraffin, 

infiltration  with 118,  226 

melting  point  of  that  used  for  embedding. . .   119 

recommended  for  cotton  plugs 99,  263 

trueing  edge  of  blocks 123 

Paraffin-oven  1 19 

Parasite, 

relation  of,  to  tissues  of  host 8, 10 

reaction  of  host  to 8 

means  of  gaining  access  to  tissues  of  host.  8,92 

Parasites,  animal,  harbored  by  plants 89 

Paratrophic  bacteria I7S 

Parenchyma,  intercellular  spaces  of,  occupied 

by  bacteria 8 

Park,  effect  of  freezing  on  B.  typhostis 247 

Pasteur, 

culture-fluid   197 

definition  of  ae'robe  and  anaerobe 230 

influence  on  bacteriology IS2 

Pathogenesis,  rules  of  proof 9 

Pathogenicity, 

developed  by  association 72, 215 

of  plant  parasites  to  animals 88 

regeneration  of 237 

Pane,  Pneumococcus,  origin  of  capsule 221 

Payen,  agar-agar,  composition  of 223 

Pear-blight  organism, 

action  of  light  on 7l 

blackening  of  foliage  attacked  by 65 

differentiated  from  B.  coli  by  KNO3 51,  113 

hosts  of 87 

natural  infection 92 

result  of  inoculations  with 202 

Pedesis i° 

Peirce,  root-tubercles  of  bur  clover 64 

Pelargonium,  natural  infection  in 92 

Pencils  for  writing  on  glass m 

Pepsin  66 


INDEX. 


28l 


Page. 

Peptone, 

commercial  45 

reaction  of  cholera  organism  with 229 

tests  for  purity  of 229 

Peptone-bacteria  175 

Peroxides,  organic,  germicidal  action  of 253 

Petri,  nitrates  in  commercial  gelatin 224 

Petri-dishes  226 

for  excluding  light 228 

for  quantitative  work  and  photographing. . .     36 

of  good  quality,  where  obtained 82 

sterilization  of 100 

Petri  und  Maassen,  hydrogen  sulphide  from 

bacteria  243 

Phenol 74 

tests  for 63 

Phenolphthalein, 

for  use  in  titration 99 

neutral  point  of 69 

Photographic  enlargements 147 

Photographic  plates,  for  special  work 136 

Photographic  prints, 

drawings  on 151 

making  of 150 

Photographing, 

of  bacteria  in  tissues 136 

of  poured-plate  colonies 134 

of  test-tube  cultures 135 

Photography, 

focusing  planars 132 

lenses  for 130, 133 

Photomicrography  263, 264 

Photomicrographic  apparatus 23, 129, 136 

care  of 139 

Photomicrographs, 

making  of 138 

time  of  exposure  for. 143 

Phragmidiothrix  162 

Physical    conditions,    influence    of,    on    mor- 
phology    177, 222 

Physiological  changes  due  to  changed  environ- 
ment        222 

Physiological   peculiarities,    for   identification 

of  species 25 

Picric  acid, 

as  a  fixing  agent 8 

germicidal  properties  of 250 

Pierce,  work  on  walnut  disease 176 

Pigment- formation, 

conditions  of 239 

discussion  of 65, 223 

effect  of  symbiosis  on 223 

how  regenerated 237 

literature  on 236 

variability  of 222, 238 

Pipettes, 

discarded,  care  of 107 

sterilization  of 100 

Pitfield's  flagella  stain 191 

Planococcus  160 

Plant  acids,  sensitiveness  of  bacteria  to 69 

Plant  juices  used  as  culture-media 41 


Page. 

Plasmolysis,  concentration  necessary  for 254 

Plastids  of  Hallier 176 

Plate-cultures, 

advantages  of,  for  isolating  bacteria n 

arrangement  of  rods  in  colonies  on 27 

characters  to  be  noted 29 

discarded,  care  of 107 

how  to  avoid  contaminations  in 103 

Koch's,  for  obtaining  pure  cultures 226 

labeling  of in 

poured,  how  made 105 

Plates, 
isochromatic,     for     photographing    stained 

sections  140 

isochromatic,  time  of  exposure  with 143 

isochromatic,  uses  of 136 

non-halation,  uses  of 136 

non-isochromatic,  when  preferable 136 

Platinum  black,  enzyme-like  action  of 234 

Platinum-iridium,     inoculating    needles     and 

loops  43 

Pleomorphism  176 

Pleuro-pneumonia,  organism  causing, 

at  limit  of  vision 18 

cultivation  of,  outside  animal  body 213 

Plugs,  paraffined 99 

Plum, 

natural  infection  in 86, 88, 92, 148 

organism  from 18 

subject  to  pear-blight 88 

Pneumococcus,  origin  of  capsule 221 

Polyangium  165 

Popoff,  germicidal  action  of  mercuric  chloride.  252 
Potassium, 

chloride,  effect  on  luminous  bacteria 60 

iodide,  effect  on  luminous  bacteria 60 

nitrate,  effect  on  luminous  bacteria 60 

nitrate,  for  differential  purposes 51 

sulfate,  effect  on  luminous  bacteria 60 

Potato,  steaming  of 98 

Potato-broth,  preparation  of 42 

Potato-starch,  aseptic,  preparation  of 50 

Prazmowski, 

arthrospores  218 

culture-fluid  197 

Precipitate,  in  culture-fluids,  nature  of 42 

Pregl,  method  of  staining 222 

Preparation  of  nutrient  agar 195 

Pressure,  effect  on  bacteria 245 

Prevention  of  disease,  methods  of 93 

Proskauer  &  Beck's  culture-fluid 198 

Prudden,  bacteria  in  ice 258 

Pseudomonas  160, 173 

Ps.  indigofera,  small  size  of 18 

Ps.  syringae,  reaction  to  nitrates 64 

Pseudo-zoogloeae  22 

Pure  cultures,  how  obtained n 

Pyocyanin,  test  for 65 

Pyoktanin  74 

Pyrogallol,  with  caustic  potash  as  absorbent  of 

oxygen 57 

Pyrogallol  developer 200, 201 


.'82 


BACTERIA   IN    RELATION    TO    PLANT    DISEASES. 


Page. 

Quantitative  determination, 

of  bacteria  in  diseased  tissues 14 

of  effect  of  freezing 79 

Quince  blight,  due  to  B.  amylovorus 202 

Rabies-virus,  removed   by  filtration 214 

Rabinowitsch,  thermophilic  bacteria 248 

Radium  rays 245 

Raulin's  culture-fluid 197 

Ravenel,  bacteria  exposed  to  liquid  air 246 

Razors, 

sharpening  of I21 

Torrey,  Rogers,  Lentz 123 

Reaction, 

for  maximum  growth  in  liquid  media. ..  69,203 
of  host  to  parasite 8 

Record-books, 

for  culture-media IO9 

for  field-notes "O 

Records,  methods  of  keeping 109 

Reducing  negatives '42 

Reducing  powers  of  bacteria 62 

Reichenbach,  branching  in  Spirillum 217 

Reichert,  thermo-regulator,  improved 78 

Reimers,  bacteria  in  soil 255 

Rein,  Florideae,  source  of  agar 224 

Reinitzer,  on  gum-ferment 233 

Re-isolations,  necessity  for 1 6 

Relationships  of  bacteria 177 

Rennet  67 

Rhabdochromatium 164 

Ribbert,  method  of  staining  capsules 194 

Rice  for  culture-media 48 

Ridgway,  color  scheme  of 263 

Roentgen  rays 245 

Rogers,  pathogenicity  developed  by  association.  215 

Rohrbeck  thermostat 75 

Roll-cultures,  Esmarch's 226 

Root-tubercle  bacteria 64, 240, 241 

Roth,  contact-irritation 253 

Rothberger, 

differential  diagnosis  with  anilin  dyes 230 

toluidin  red  for  differentiation 230 

Rothert,  effect  of  ether  and  chloroform 254 

Roux  filter,  for  culture-fluids,  etc 44 

Roux  thermo-regulator 78 

Rubber, 

caps  for  tubes  of  media 99 

note  on  best  solvents  for 207 

Rulers  for  measuring  colonies,  glass,  steel. ...   115 

Rules  of  proof 9,  10 

Russell  &  Babcock,  fermentation  of  silage. . . .  257 

Ruzicka,  inner  structure  of  bacteria 217 

Saccardo,  color  scheme  of 263 

Sachs,  reduction  of  cane-sugar  in  plants 66 

Safranin-picro-nigrosin  189 

Salicylic  acid 74 

Salt-bouillon 51 

Salt-water    bacteria,    minimum    temperature 

for 87, 255 

Sander,  growth  of  tubercle  bacteria  on  vege- 
table media 222 

Sand-fleas,  bacterium  of  luminous 60, 242 


Page. 

Saponification  of  casein  in  milk-cultures 46 

Saprophytes, 

behavior  of,  when  injected  into  plants 89 

presence  in  diseased  parts  long  affected ....     13 

Sarcina 160 

Sauerkraut,  fermentation  of 257, 258 

Scales  for  photographic  exposures 143 

Schaffner's  safranin-picro-nigrosin 189 

Schaudinn, 

bacillus  of  large  size 19 

two  endospores  in  a  cell 22 

Scheffler,  neutral  red  for  differentiating 230 

Schild,  formalin  for  detection  of  B.  typhosus. .  229 
Schill,  preparation  of  cultures  for  museums. .  262 
Schilow,   germicidal   value   of   hydrogen   pe- 
roxide   251 

Schneider,  studies  on  pigment- formation 237 

Schottelius, 

B.  prodigiosus,  non-pigmented  races  of 222 

preparation  of  nutrient  agar 223 

Schliiter,  effect  of  acids  on  bacteria 249 

Schultz,  structural  changes  due  to  antiseptics.  252 

Schumburg,  bromine,  water  treated  with 252 

Schu'tz,  method  of  making  nutrient  agar 34 

Schwartz,  on  antiseptics 250 

Sclavo's  flagella-stain 192 

Sclerothrix  172 

Sea-weeds  furnishing  agar 33 

Sections,  microtome, 

balsam  for  mounting 117 

boxes  for  preservation  of 117 

cutting  and  care  of 119 

for  photographing 140 

keeping  of  material  for 117 

mounting  of 119 

preparation  of 1 18 

staining  of 119 

Sedgwick  &   Winslow,   influence  of  cold  on 

B.  typhosus 247 

Setchell,  thermophilic  bacteria 248 

Signs  of  disease 7 

Silage-fermentation, 

bacteria  active  in 256 

bacteria  not  active  in 257 

Silicate-jelly, 

for  differential  purposes 39 

method  of  preparing 37,  206 

value,  as  a  culture-medium 36 

Winogradsky^Sleskin  198 

Sjoebring,  nucleus  in  bacteria 216 

Skschivan,  branched  forms 217 

Slater,  branched  forms 237 

Slide-boxes 117 

Slides, 

care  of  discarded 107 

numbering  of T2I 

Smith  &  Swingle,  effect  of  freezing  on  bac- 
teria       83 

Smith,  Theobald, 

discovers  cause  of  Texas  fever 153 

method  of  filtering 226 

tubercle  bacteria  difficult  to  destroy  by  heat.  247 
Soaps,  germicidal  action  of  certain 253 


INDEX. 


283 


Pago. 
Sodium  chloride, 

effect  on  luminous  bacteria 60 

restraining  influence  of 70,  252 

Sodium  fluoride,  germicidal  action  of 253 

Sodium  hydrate,  method  of  increasing  tolera- 
tion of 70 

Sodium  nitrite,  used  for  indol  test 62 

Softening  hard  tissues,  fluid  for 200 

Soft-rot  bacteria,  many  plants  attacked  by. ...     87 
Soil, 

bacteria  in  upper  layers  of 255 

parasites  carried  in 91 

sterilization  of 85 

Solid  vegetable  media, 

behavior  of  organisms  on 40 

preparation  of 40 

substances  recommended 40 

Solio  paper,  toning  bath  for 201 

Solubility  of  glassware 129, 223 

Solution  of  tissues  occupied  by  bacteria 10 

Solution  persulphate  of  iron,  preparation  of. .   188 

Sorghum,  natural  infection  in 92 

Soy-bean,  natural  infection  in 92 

Sphaerotilus    162 

Species, 

morphology  not  sufficient  for  differentiation .     25 
Specimens,  method  of  substituting  alcohol  for 

water  in 226 

Spectrum,  part  possessing  germicidal  action. .  244 

Spencer  microscopes 129 

Spina,  studies  of  reduction  processes 239 

Spirillum  161 

branching  in 217 

double  staining  of 217 

Ehrenberg's  genus 173 

Spirillum  undula, 

action  of  ether  on 254 

flagella  of 20 

Spirillum  volutans, 

flagella  of 20 

reaction  to  stimuli 27 

Spirochaeta  lot 

Spirodiscus  162 

Spiromonas  162 

Spirosoma  161 

Spores, 

action  of  heat  on 84 

Aujeszky's  stain 219 

bacilli  bearing  green 218 

cause  of  resistance  to  dry  heat 218 

classification  based  on 157 

common  in  stringy  bread 261 

effect  of  steam  on 246 

Fiocca's  stain 218 

Foth's  stain 218 

germination  of 21 

influence    of    environment    on     formation 

Of 219,  222 

Klein's  stain 219 

means  for  identification  of 21 

Moeller's  stain 218 

reaction  to  stains 27 


Page. 

Spores — continued, 
resistance  to  high  temperatures  and  steam 

heat  21, 84, 246 

resistant,  present  in  meat-extracts 260 

resistant,  present  in  milk 46 

stains  for. 194 

Spraying,  inoculation  by 108 

Stadler,  effect  of  sodium  chloride  on  bacteria.  252 

Stage-micrometer  115 

Staining, 

bacteria  in  tissues 29, 222 

double 217,218 

microtome-sections  119 

Staining-media,  groups  of 28 

Staining  methods 221 

bacteria,  vegetative  forms 27,187 

capsules  19,  194 

flagella  20, 21, 27,  189 

spores  27, 194 

tissues 187 

Staining  produced  by  bacteria, 

in  host-plant 65 

in  nutrient  substrata 65, 211, 214, 237 

Stains, 

anilin,  alcoholic  solutions  of 187 

general  and  miscellaneous 187 

Starch  from  potato,  preparation  of 50 

Starch-jelly,  nutrient 50,  196 

Starch,  test  for  slime  derived  from 221 

Starrett  cross-level ^9 

Starvation,  resistance  to 223 

Statistics,  need  of,  on  plant  diseases 90 

Steam, 

spores  resistant  to 84 

sterilization  by,  possible  source  of  error  in.     47 

vegetative  bacteria  sensitive  to 83 

Steaming, 

media  in  tubes 48 

milk  for  culture-media 46, 47 

Steam-sterilizer,  Arnold 47 

Stephens,  flagella-stain 220 

Sterilization, 

cold,  of  culture-fluids 43, 52 

cotton   IOI 

culture-media  85 

dry,  of  pipettes,  scalpels,  etc 41 

glassware,  instruments 100, 107 

infectious  material '  IO6 

milk  46 

oven  for loo 

silicate-jelly  JQ 

soils   \]       8s 

solid  vegetable  media 40 

starch-jelly gO 

surface   of  plants   before   making   cultures 

from j-j 

surface  of  plants  to  be  inoculated 41, 108 

syringes   IO2 

with  metallic  copper 97  2r? 

Still, 

for  water,  in  small  quantities 128 

for  water,  on  a  large  scale 124 


284 


BACTERIA    IN    RELATION    TO    PLANT   DISEASES. 


Page. 

Stock-cultures,  how  best  kept 72, 123 

Stoddart,  media  for  differentiation  of  motile 

bacteria  2(> 

Stolz,  peculiar  growths  in  Pneumococcus,  etc.  217 
Stomata, 

artificial  infections  by  way  of 108, 126 

natural  infection  through 9°, 92 

Stored  media,  effect  of  loss  of  water  on 99 

Streak-cultures,  characters  to  be  noted 3° 

Streblothrichia   l62 

Streptococcus   • • Io° 

Streptococcus  mesenterioides,  isolation  of  by 

heat  Io6 

Streptothrix Io2 

Succession  of  organisms  in  diseased  tissues..     73 

Sugar-cane,  red  stain  in  bundles  of 66 

Sugars, 

effect  on  liquefaction  of  gelatin 29 

growth  retarded  by 223, 248 

proper  sterilization  of  media  containing 99 

reagents  for 208, 234 

Sulphur-bacteria  162, 261 

Sunlight, 

effect  of 71 

in  photomicrography 24 

Surface  organisms,  partial  removal  by  washing .     14 

Surface  sterilization !3 

Synthetic  culture-media, 

kinds  of 49, 197 

value  of Si 

Syringes,  hypodermic I02 

Systematic  position  of  bacteria 177 

Szyszylowicz,  corallin  as  a  microchemical  re- 
agent        221 

Tannin,  method  for  detecting  in  cells 207 

Temperature, 

of  liquid  air •  83, 247 

optimum   75 

maximum   75 

minimum   76 

range  of,  suited  to  growth 86 

Temperature,  influence  on, 

geographical  distribution 7 

involution-forms  22 

Teratological  growths 23 

Test-tube  cultures, 

inoculation  of 105 

labeling  of m 

photographing  of 135 

Test-tubes, 

device  for  filling  rapidly 98, 227 

holder  for 104 

Jena  glass 81 

resistant  glass 82 

tests  of  solubility 129 

Tetanus  poison,  lethal  dose  of 213 

Thaxter,  Myxobacteria 164 

Thermal  death-point, 

apparatus  for 78 

method  of  determining 77 

range  of 87 


Page. 
Thermal  death-point  experiments,  methods  of 

checking  185 

Thermal  limits,  effect  of,  on  pigment  produc- 
tion, pathogenicity,  sporulation 87 

Thermal  relations, 

of  bacteria 75 

of  enzymes 67 

Thermometers,  Anschiitz  normal 78 

Thermophilic  bacteria, 

common  in  digestive-tract 248 

literature  on 247 

maximum  temperature  for 87 

minimum  temperature  for 87 

often  spore-bearing 248 

Thermo-regulator, 

Novy  228 

Reichert  78 

Roux   78 

Tollens   37 

Thermostat-room  75 

Thermostats  75 

Thermotaxis  254 

Thiocapsa 163 

Thiocystis  163 

Thiodictyon   164 

Thiopedia  163 

Thiopolycoccus   164 

Thiosarcina  163 

Thiospirillum  164 

Thiothece   164 

Thiothrix   162 

Thomann,  bacterium  of  stringy  bread 261 

Thumm,  green-fluorescent  bacteria 238 

Thymol   74 

Tischutkin,  preparation  of  nutrient  agar 224 

Tissues,  solution  of,  by  bacteria 10 

Titration  of  culture-media 69, 99 

Tobacco,  fermentation  of 257 

Tollens,  thermo-regulator 37 

Toluidin  red,  for  differentiation 230 

Toning  bath  for  solio  paper 201 

Transfers,  how   to   avoid   contaminations   in 

making  103 

Trenkmann,  flagella-stain 219 

Trevisan,  genus   Pacinia 172 

Trikresol   74 

Trypsin   66 

Tsiklinsky,  thermophilic  bacteria 248 

Tubes, 

care  of  discarded 107 

rapid  filling,  with  fluid  media 99 

Tumors  produced  by  bacteria 8 

Typhoid  bacillus,  effect  of  copper  on. .  74,250,253 

Tyrosin,  qualitative  test  for 63 

Ultramicroscopical  organisms 18, 211, 213 

Unguentum  resinae 228 

Urocephalum  159, 264 

Uschinsky's  solution, 

cultures  in  dilute,  for  flagella  staining 20 

formula  197 

in  silicate-jelly 39 

modified 197 


INDEX. 


285 


Page. 

U-tubes,  for  testing  growth  in  nitrogen 58 

Vacuo, 

concentration  of  fluids  in 68 

growth  in 54 

Vallin,  cleaning  filters 227 

Van  Ermengem, 

flagella-stain rgi 

germicidal  effect  of  ozone 251 

van't  Hoff,  modification  of  plate-method 228 

van  Tieghem,  thermophilic  bacteria 247 

Varietal  resistance  to  disease 93, 186 

Vascular  system  occupied  by  bacteria 8 

Velox-prints   151 

Ventilation,  of  dark-room 149 

Vernhout,  bacterial  fermentation  of  tobacco. .  257 

Vibrio,  use  of,  as  a  genus  name 173 

Vibrio  cholerae, 

action  of  copper  on 74, 250 

classification   172 

Vibrio,  Muller's  genus 172 

Viscidity  42 

of  bacterial  growths 19 

of  milk 46 

of  Uschinsky's  solution 20 

Vitality  on  various  media 72 

Voges, 

blue  water-bacteria 237 

cultivates  Cholera  vibrio  on  potato 249 

Voigtlaender,  physical  properties  of  agar 224 

von  Esmarch,  roll-cultures  of 226 

von  Freudenreich,  filtration  of  agar 224 

von  Rozsahegyi,  pigmented-media  for  differ- 
entiation   229 

V-shaped  forms 23,  217 

Wager  exposure  scale 143 

Waldo  &  Walsh,  bread  not  fully  sterilized  by 

baking   261 

Wall-charts    206 

Wall  of  cell,  outer,  reaction  to  stains 28 

Walliczek, 

effect  of  dry  air  on  bacteria 249 

effect  of  tannin  on  bacteria 251 

Ward,  A.  R.,  ropy  milk 259,  260 

Ward,  H.  M.,  bacteria  from  Thames  water.. .  258 

Warming  20 

Water, 

apparatus  for  distilling 124 

bacteria  in,  literature  on 258 

Water-bacteria, 

blue  and  violet 237 

Hesse  &  Niedner's  nutrient  agar  for 196 


Page. 
Water-bacteria — continued. 

how  best  kept 263 

media  for 258 

Water,  examination  of.  English  methods  for.  220 

Water-bath,  Ostwald-Pfeffer 78 

Water-pores,  infection  through 92, 102, 124 

Wax-mixture,  Darwin's 200 

Wehmer,  fermentation  of  sauerkraut 2=;8 

Weigert,  early  use  of  anilin  stains 29 

Weiss, 

bacteria  from  soured  foods 235 

ruled  counting-plate 36 

Welch, 

capsule-stain  20, 194 

influence  of 152 

Welcke,  flagella-stain 220 

Wescner,  eggs  as  a  culture-medium 225 

Weyl,  ozone  as  a  germicide 252 

Wilhelmy,  bacteria  in  meat-extracts 260 

Williams,  flagella-stain 193 

Winogradsky, 

agar  for  isolation  of  nitrate  bacteria 199 

elective  cultures 106 

medium  for  nitrogen-assimilating  bacteria.   199 

nitrifying  bacteria 240 

retting  of  flax 257 

silicate-jelly  36.-24O 

red  sulphur-bacteria 263 

Winogradsky  &  Omelianski,  fluid-media  for 

isolating  nitrate  and  nitrite  bacteria. . .   199 

Winogradsky-Sleskin  silicate-jelly 108 

Winslow  &  Rogers,  Coccaceae  revised  by 265 

Wood-vinegar,  an  energetic  disinfectant 252 

Wounds, 

disinfection  of 107 

natural  infection  through 92 

Wright,  anaerobes,  simple  methods  of  culti- 
vating     228 

Wynne  exposure  meter 143 

Yendo,  source  of  commercial  agar 225 

Yersen,  pest  carried  by  rats 215 

Yokote,  filtration  of  agar 225 

Y-shaped  forms 23 

Zeiss  lenses  for  photographic  work, 

Double-Protar  130 

Planar  132 

Unar   130 

Zeiss  microscopes 129, 130 

Zettnow, 

cleaning  cover-glasses 227 

flagella-stain   192 

light-filter 137, 201 


M 


